U.S. patent number 7,550,501 [Application Number 11/697,651] was granted by the patent office on 2009-06-23 for compounds and methods for treating toll-like receptor 2-related diseases and conditions.
This patent grant is currently assigned to Eisai R&D Management Co., Ltd.. Invention is credited to Jesse Chow, Fabian Gusovksy, Lynn Hawkins, Mark Spyvee.
United States Patent |
7,550,501 |
Chow , et al. |
June 23, 2009 |
Compounds and methods for treating toll-like receptor 2-related
diseases and conditions
Abstract
The present invention relates to compounds and methods useful in
the prevention or treatment of diseases or conditions associated
with Toll-like receptor 2 activation.
Inventors: |
Chow; Jesse (Hooksett, NH),
Gusovksy; Fabian (Andover, MA), Hawkins; Lynn (Concord,
MA), Spyvee; Mark (Hampstead, NH) |
Assignee: |
Eisai R&D Management Co.,
Ltd. (Tokyo, JP)
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Family
ID: |
34520187 |
Appl.
No.: |
11/697,651 |
Filed: |
April 6, 2007 |
Prior Publication Data
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Document
Identifier |
Publication Date |
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US 20070167409 A1 |
Jul 19, 2007 |
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Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
Issue Date |
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10973164 |
Oct 25, 2004 |
7202234 |
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60514283 |
Oct 24, 2003 |
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Current U.S.
Class: |
514/419;
514/80 |
Current CPC
Class: |
A61P
31/04 (20180101); A61P 1/04 (20180101); A61P
43/00 (20180101); A61P 17/10 (20180101); A61P
41/00 (20180101); A61K 31/4045 (20130101); A61P
37/00 (20180101); A61P 37/08 (20180101); C07F
9/02 (20130101); A61P 19/02 (20180101); A61P
1/02 (20180101); A61P 37/02 (20180101); A61P
9/10 (20180101); C07F 9/091 (20130101); A61P
31/12 (20180101); A61P 17/00 (20180101); A61P
11/06 (20180101); A61P 29/00 (20180101); A61P
17/02 (20180101); A61P 25/28 (20180101); A61P
31/00 (20180101); A61P 33/02 (20180101); A61P
25/00 (20180101); A61P 31/22 (20180101); A61P
17/06 (20180101); C07F 9/5728 (20130101); A61P
9/00 (20180101); A61P 11/00 (20180101); A61P
31/08 (20180101); A61K 31/661 (20130101); A61P
13/12 (20180101) |
Current International
Class: |
A61K
31/661 (20060101); A61K 31/4045 (20060101) |
Field of
Search: |
;514/80,419 ;548/414
;568/14 ;558/169 |
References Cited
[Referenced By]
U.S. Patent Documents
Foreign Patent Documents
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1 147 117 |
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Jun 2004 |
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EP |
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WO 98/50547 |
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Nov 1998 |
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WO |
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WO 03/020760 |
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Mar 2003 |
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WO |
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Primary Examiner: Anderson; Rebecca L
Assistant Examiner: Nolan; Jason
Attorney, Agent or Firm: Clark & Elbing LLP
Parent Case Text
CROSS-REFERENCE TO RELATED APPLICATIONS
This application is a divisional of, and claims priority from, U.S.
patent application Ser. No. 10/973,164, filed Oct. 25, 2004 (U.S.
Pat. No. 7,202,234), which claims the benefit of the filing date of
U.S. provisional patent application No. 60/514,283, filed Oct. 24,
2003, the contents of which are incorporated by reference herein in
their entirety.
Claims
What is claimed is:
1. A method of treating a disease or condition characterized by
Toll-like receptor 2 activation in a patient, wherein said disease
or condition is selected from the group consisting of inflammatory
bowel disease, sepsis, periodontal disease, acne, chronic
obstructive pulmonary disease, arthritis, cystic fibrosis,
ischemia/reperfusion injury, asthma, necrotizing enterocolitis,
leprosy, atopic dermatitis, psoriasis, allergies, coronary artery
bypass grafting, and atherosclerosis said method comprising
administering to the patient a Toll-like receptor 2 antagonist of
formula I ##STR00254## or a pharmaceutically acceptable salt
thereof, wherein a is an integer of 1 to 3; b is an integer of 0 to
4, wherein when b is 0, the carbon bonded to X and W is not bonded
to 2 or more heteroatoms; R.sup.1 is H or C.sub.1-6 alkyl; X is
selected from the group consisting of --NR.sup.X1V,
--N(R.sup.X1)C(O)V, --N(R.sup.X1)C(S)V,
--N(R.sup.X1)C(O)N(R.sup.X2)V, --N(R.sup.X1)C(S)N(R.sup.X2)V,
--N(R.sup.X1)C(O)OV, --N(R.sup.X1)S(O).sub.2V, --C(O)N(R.sup.X1)V,
--C(O)OV, --OC(O)V, --OC(O)OV, and --OC(O)N(R.sup.X1)V, where each
of R.sup.X1 and R.sup.X2 is, independently, H or C.sub.1-6 alkyl,
and V is a C.sub.1-20 alkyl, C.sub.1-20 alkenyl, or C.sub.1-20
alkynyl group, optionally substituted with halo, hydroxyl,
C.sub.1-21 acyloxy, oxo, C.sub.1-20 alkoxyl, or C.sub.1-20
thioalkoxyl and optionally containing 1 or 2 phenyl or biphenyl
moieties in and/or at the end of the carbon chain; W is selected
from the group consisting of H, --C(O)N(R.sup.W1)R.sup.W2,
--C(O)OR.sup.W2, --(CH.sub.2).sub.cOR.sup.W3,
--(CH.sub.2).sub.cSR.sup.W3,
--(CH.sub.2).sub.cO(CH.sub.2).sub.dCH(OR.sup.W3)R.sup.W4,
--(CH.sub.2).sub.cS(CH.sub.2).sub.dCH(OR.sup.W3)R.sup.W4,
--C(O)N(R.sup.W1)(CH.sub.2).sub.cCH(OR.sup.W3)R.sup.W4, and
--C(O)N(R.sup.W1)(CH.sub.2).sub.cCH(OR.sup.W3)(CH.sub.2).sub.eOR.sup.W5,
wherein each of c and d is, independently, an integer of 1 to 4, e
is an integer of 2 to 4, R.sup.W1 is H or C.sub.1-6 alkyl, R.sup.W2
is C.sub.1-20 alkyl, C.sub.1-20 alkenyl, or C.sub.1-20 alkynyl,
each of R.sup.W3 and R.sup.W5 is, independently, H, C.sub.1-20
alkyl, C.sub.1-21 acyl, C.sub.1-20 alkenyl, or C.sub.1-20 alkynyl,
and R.sup.W4 is H, C.sub.1-20 alkyl, C.sub.1-20 alkenyl, or
C.sub.1-20 alkynyl, wherein each of R.sup.W2, R.sup.W3, R.sup.W4,
and R.sup.W5 is optionally substituted with halo, hydroxyl,
C.sub.1-21 acyloxy, oxo, C.sub.1-20 alkoxyl, or C.sub.1-20
thioalkoxyl, optionally contains 1 to 2 phenyl or biphenyl moieties
in and/or at the end of the carbon chain, and optionally contains 1
to 4 non-vicinal oxygen atoms in the carbon chain; and U is
selected from the group consisting of ##STR00255## wherein f is an
integer of 1 to 4, g is 0 or 1, each of R.sup.U1, R.sup.U2, and
R.sup.U3 is, independently, H, optionally substituted C.sub.1-6
alkyl, optionally substituted C.sub.7-16 aralkyl, or optionally
substituted C.sub.2-15 heterocyclylalkyl, or R.sup.U1 is H or
optionally substituted C.sub.1-6 alkyl and R.sup.U2 and R.sup.U3
together with the carbon atom they are bonded to form an optionally
substituted C.sub.3-6 aliphatic ring, or R.sup.U2 is H and R.sup.U3
and R.sup.U1 together with the carbon atom bonded to R.sup.U3 and
the nitrogen atom bonded to R.sup.U1 form an optionally substituted
4-6- membered heterocyclic ring, and R.sup.U4 is selected from the
group consisting of --C(O)R.sup.U5 and --C(O)OR.sup.U6 wherein
R.sup.U5 is an optionally substituted C.sub.1-9 indole, and
R.sup.U6 is an optionally substituted C.sub.1-6 alkyl.
2. The method of claim 1, wherein X or W contains at least one
linear alkyl moiety of 7 or more carbons.
3. The method of claim 1, wherein each of X and W contains at least
one linear alkyl moiety of 7 or more carbons.
4. The method of claim 1, wherein W is
--C(O)NH(CH.sub.2).sub.2CH(OH)R.sup.W4, wherein R.sup.W4 is
C.sub.7-19 alkyl.
5. The method of claim 1, wherein W is
--C(O)NH(CH.sub.2).sub.2CH.sub.2OR.sup.W3, wherein R.sup.W3 is
--C(O)(CH.sub.2).sub.aaCH.sub.3, wherein aa is an integer of 6 to
18.
6. The method of claim 1, wherein W is
--C(O)NH(CH.sub.2).sub.2CH(OR.sup.W3)R.sup.W4, wherein R.sup.W3 is
--C(O)(CH.sub.2).sub.aaCH.sub.3 and R.sup.W4 is
CH.sub.2OC(O)(CH.sub.2).sub.bbCH.sub.3, wherein each of aa and bb
is, independently, an integer of 6 to 18.
7. The method of claim 1, wherein U is
--C(O)C(R.sup.U2)(R.sup.U3)NHR.sup.U4 or
--C(O)(CH.sub.2).sub.fNHR.sup.U4 or wherein R.sup.U2 is an
optionally substituted C.sub.1-6 alkyl, R.sup.13 is H, and R.sup.U4
--C(O)R.sup.U5, wherein R.sup.U5 is an optionally substituted
C.sub.2-9 indole, or an optionally substituted C.sub.2-9
heteroaryl.
8. The method of claim 1, wherein U is ##STR00256## wherein
R.sup.U2 is C.sub.1-6 alkyl, R.sup.U3 is H, and R.sup.U12 is
optionally substituted C.sub.6-10 aryl, optionally substituted
C.sub.6-10 aryloxy, optionally substituted C.sub.7-16 aralkyl,
optionally substituted C.sub.7-16 aralkoxy, optionally substituted
C.sub.2-9 heterocyclyl, optionally substituted C.sub.2-9
heterocyclyloxy, optionally substituted C.sub.3-15
heterocyclylalkyl, or optionally substituted C.sub.3-15
heterocyclylalkyloxy.
9. The method of claim 1, wherein U is selected from the group
consisting of ##STR00257##
10. The method of claim 1, wherein: X is selected from the group
consisting of --NR.sup.X1V, --N(R.sup.X1)C(O)V, --N(R.sup.X1)C(S)V,
--N(R.sup.X1)C(O)N(R.sup.X2)V, --N(R.sup.X1)C(S)N(R.sup.X2)V,
--N(R.sup.X1)C(O)OV, --N(R.sup.X1)S(O).sub.2V, --C(O)N(R.sup.X1)V,
--C(O)OV, --OC(O)OV, and --OC(O)N(R.sup.X1)V, where each of
R.sup.X1 and R.sup.X2 is, independently, H or C.sub.1-6 alkyl, and
V is a C.sub.1-20 alkyl, C.sub.1-20 alkenyl, or C.sub.1-20 alkynyl
group, optionally substituted with halo, hydroxyl, C.sub.1-21
acyloxy, oxo, C.sub.1-20 alkoxyl, or C.sub.1-20 thioalkoxyl and
optionally containing 1 or 2 phenyl or biphenyl moieties in and/or
at the end of the carbon chain.
11. The method of claim 1, wherein: W is selected from the group
consisting of H, --C(O)N(R.sup.W1)R.sup.W2, --C(O)OR.sup.W2,
--(CH.sub.2).sub.cSR.sup.W3,
--(CH.sub.2).sub.cO(CH.sub.2).sub.dCH(OR.sup.W3)R.sup.W4,
--(CH.sub.2).sub.CS(CH.sub.2).sub.dCH(OR .sup.W3)R.sup.W4,
--C(O)N(R.sup.W1)(CH.sub.2).sub.cCH(OR.sup.W3)R.sup.W4, and
--C(O)N(R.sup.W1)(CH.sub.2).sub.cCH(OR.sup.W3)(CH.sub.2).sub.eOR.sup.W5,
wherein each of c and d is, independently, an integer of 1 to 4, e
is an integer of 2 to 4, R.sup.W1 is H or C.sub.1-6alkyl, R.sup.W2
is C.sub.1-20 alkyl, C.sub.1-20 alkenyl, or C.sub.1-20 alkynyl,
each of R.sup.W3 and R.sup.W5 is, independently, H,
C.sub.1-20alkyl, C.sub.1-21acyl, C.sub.1-20alkenyl, or
C.sub.1-20alkynyl, and R.sup.W4 is H, C.sub.1-20 alkyl, C.sub.1-20
alkenyl, or C.sub.1-20 alkynyl, wherein each of R.sup.W2, R.sup.W3,
R.sup.W4,and R.sup.W5 is optionally substituted with halo,
hydroxyl, C.sub.1-21 acyloxy, oxo, C.sub.1-20 alkoxyl, or
C.sub.1-20 thioalkoxyl, optionally contains 1 to 2 phenyl or
biphenyl moieties in and/or at the end of the carbon chain, and
optionally contains 1 to 4 non-vicinal oxygen atoms in the carbon
chain.
12. The method of claim 1, wherein: X is selected from the group
consisting of --NR.sup.X1V, --N(R.sup.X1)C(O)V, --N(R.sup.X1)C(S)V,
--N(R.sup.X1)C(O)N(R.sup.X2)V, --N(R.sup.X1)C(S)N(R.sup.X2)V,
--N(R.sup.X1)C(O)OV, --N(R.sup.X1)S(O).sub.2V, --C(O)N(R.sup.X1)V,
--C(O)OV, --OC(O)OV, and --OC(O)N(R.sup.X1)V, where each of
R.sup.X1 and R.sup.X2 is, independently, H or C.sub.1-6 alkyl, and
V is a C.sub.1-20 alkyl, C.sub.1-20 alkenyl, or C.sub.1-20alkynyl
group, optionally substituted with halo, hydroxyl, C.sub.1-21
acyloxy, oxo, C.sub.1-20 alkoxyl, or C.sub.1-20 thioalkoxyl and
optionally containing 1 or 2 phenyl or biphenyl moieties in and/or
at the end of the carbon chain; and W is selected from the group
consisting of H, --C(O)N(R.sup.W1)R.sup.W2, --C(O)OR.sup.W2,
--(CH.sub.2).sub.cSR.sup.W3,
--(CH.sub.2).sub.cO(CH.sub.2).sub.dCH(OR.sup.W3)R.sup.W4,
--(CH.sub.2).sub.cS(CH.sub.2).sub.dCH(OR.sup.W3)R.sup.W4,
--C(O)N(R.sup.W1)(CH.sub.2).sub.cCH(OR.sup.W3)R.sup.W4, and
--C(O)N(R.sup.W1)(CH.sub.2).sub.cCH(OR.sup.W3)(CH.sub.2).sub.eOR.sup.W5,
wherein each of c and d is, independently, an integer of 1 to 4, e
is an integer of 2 to 4, R.sup.W1 is H or C.sub.1-6alkyl, R.sup.W2
is C.sub.1-20 alkyl, C.sub.1-20 alkenyl, or C.sub.1-20 alkynyl,
each of R.sup.W3 and R.sup.W5 is, independently, H, C.sub.1-20
alkyl, C.sub.1-21 acyl,C.sub.1-20 alkenyl, or C.sub.1-20
alkynyl,and R.sup.W4 is H, C.sub.1-20 alkyl, C.sub.1-20 alkenyl, or
C.sub.1-20 alkynyl, wherein each of R.sup.W2, R.sup.W3, R.sup.W4,
and R.sup.W5 is optionally substituted with halo, hydroxyl,
C.sub.1-21 acyloxy, oxo, C.sub.1-20 alkoxyl, or C.sub.1-20
thioalkoxyl, optionally contains 1 to 2 phenyl or biphenyl moieties
in and/or at the end of the carbon chain, and optionally contains 1
to 4 non-vicinal oxygen atoms in the carbon chain.
13. The method of claim 1, wherein: X is selected from the group
consisting of --N(R.sup.X1)C(O)V, --N(R.sup.X1)C(S)V,
--N(R.sup.X1)C(O)N(R.sup.X2)V, --N(R.sup.X1)C(S)N(R.sup.X2)V,
--N(R.sup.X1)C(O)OV, and --N(R.sup.X1)S(O).sub.2V, where each of
R.sup.X1 and R.sup.X2 is, independently, H or C.sub.1-6 alkyl, and
V is a C.sub.1-20 alkyl, C.sub.1-20 alkenyl, or C.sub.1-.sub.1-20
alkynyl group, optionally substituted with halo, hydroxyl,
C.sub.1-21 acyloxy, oxo, C.sub.1-20 alkoxyl, or C.sub.1-20
thioalkoxyl and optionally containing 1 or 2 phenyl or biphenyl
moieties in and/or at the end of the carbon chain.
14. The method of claim 1, wherein: W is selected from the group
consisting of --C(O)N(R.sup.W1)R.sup.W2,
--(CH.sub.2).sub.cSR.sup.W3,
--(CH.sub.2).sub.cO(CH.sub.2).sub.dCH(OR.sup.W3)R.sup.W4,
--(CH.sub.2).sub.cS(CH.sub.2).sub.dCH(OR.sup.W3)R.sup.W4,
--C(O)N(R.sup.W1)(CH.sub.2).sub.cCH(OR.sup.W3)R.sup.W4, and
--C(O)N(R.sup.W1)(CH.sub.2).sub.cCH(OR.sup.W3)(CH.sub.2).sub.eOR.sup.W5,
wherein each of c and d is, independently, an integer of 1 to 4, e
is an integer of 2 to 4, R.sup.W1 is H or C.sub.1-6 alkyl,
R.sup.W2is C.sub.1-20 alkyl, C.sub.1-20 alkenyl, or C.sub.1-20
alkynyl, each of R.sup.W3 and R.sup.W5 is, independently, H,
C.sub.1-20 alkyl, C.sub.1-21 acyl, C.sub.1-20 alkenyl, or
C.sub.1-20 alkynyl, and R.sup.W4 is H, C.sub.1-20 alkyl, C.sub.1-20
alkenyl, or C.sub.1-20 alkynyl, wherein each of R.sup.W2, R.sup.W3,
R.sup.W4 and R.sup.W5 is optionally substituted with halo,
hydroxyl, C.sub.1-21 acyloxy, oxo, C.sub.1-20 alkoxyl, or
C.sub.1-20 thioalkoxyl, optionally contains 1 to 2 phenyl or
biphenyl moieties in and/or at the end of the carbon chain, and
optionally contains 1 to 4 non-vicinal oxygen atoms in the carbon
chain.
15. The method of claim 1, wherein said compound is ##STR00258## or
a pharmaceutically acceptable salt thereof.
16. The method of claim 1, wherein said compound is ##STR00259## or
a pharmaceutically acceptable salt thereof.
17. The method of claim 1, wherein said compound is ##STR00260## or
a pharmaceutically acceptable salt thereof.
18. The method of claim 1, wherein said compound is ##STR00261## or
a pharmaceutically acceptable salt thereof.
19. The method of claim 1, wherein said compound is ##STR00262## or
a pharmaceutically acceptable salt thereof.
20. The method of claim 1, wherein said compound is ##STR00263## or
a pharmaceutically acceptable salt thereof.
21. The method of claim 1, wherein said compound is ##STR00264## or
a pharmaceutically acceptable salt thereof.
22. The method of claim 1, wherein said compound is ##STR00265## or
a pharmaceutically acceptable salt thereof.
23. The method of claim 1, wherein said compound is ##STR00266## or
a pharmaceutically acceptable salt thereof.
24. The method of claim 1, wherein said compound is ##STR00267## or
a pharmaceutically acceptable salt thereof.
25. The method of claim 1, wherein said compound is ##STR00268## or
a pharmaceutically acceptable salt thereof.
26. The method of claim 1, wherein said compound is ##STR00269## or
a pharmaceutically acceptable salt thereof.
27. The method of claim 1, wherein said compound is ##STR00270## or
a pharmaceutically acceptable salt thereof.
28. The method of claim 1, wherein said compound is ##STR00271## or
a pharmaceutically acceptable salt thereof.
29. The method of claim 1, wherein said compound is ##STR00272## or
a pharmaceutically acceptable salt thereof.
30. The method of claim 1, wherein said compound is ##STR00273## or
a pharmaceutically acceptable salt thereof.
31. The method of claim 1, wherein said compound is ##STR00274## or
a pharmaceutically acceptable salt thereof.
32. The method of claim 1, wherein said compound is ##STR00275## or
a pharmaceutically acceptable salt thereof.
33. The method of claim 1, wherein said compound is ##STR00276## or
a pharmaceutically acceptable salt thereof.
34. The method of claim 1, wherein said compound is ##STR00277## or
a pharmaceutically acceptable salt thereof.
Description
FIELD OF THE INVENTION
This invention relates to the prevention and treatment of diseases
and conditions associated with Toll-like receptor 2 activation.
BACKGROUND OF THE INVENTION
The vertebrate immune system protects the body against undesirable
foreign matter that enters the body, such as infecting pathogens
(e.g., bacteria, viruses, fungi, and parasites) and their
by-products. One manner by which this takes place involves the
adaptive immune system, through which the body recognizes foreign
antigens and generates specific immune responses against them. The
induction of adaptive immunity takes time (e.g., 2-3 days post
infection), and thus could leave the body vulnerable to the adverse
effects of early infection, if it were not for the action of
another division of the immune system, the innate immune
system.
The innate immune system provides the body with a first line
defense against invading pathogens. In an innate immune response,
an invading pathogen is recognized by a germline-encoded receptor,
the activation of which initiates a signaling cascade that leads to
the induction of cytokine expression. Innate immune system
receptors have broad specificity, recognizing molecular structures
that are highly conserved among different pathogens. These
receptors are known as Toll-like receptors (TLRs), due to their
homology with receptors that were first identified and named in
Drosophila, and are present in cells such as macrophages, dendritic
cells, and epithelial cells.
There are at least ten different TLRs in mammals, and ligands and
corresponding signaling cascades have been identified for some of
these receptors. For example, TLR2 is activated by the lipoprotein
of bacteria (e.g., E. coli), TLR3 is activated by double-stranded
RNA, TLR4 is activated by lipopolysaccharide (i.e., LPS or
endotoxin) of Gram-negative bacteria (e.g., Salmonella and E. coli
O157:H7), TLR5 is activated by flagellin of motile bacteria (e.g.,
Listeria), and TLR9 is activated by unmethylated CpG sequences of
pathogen DNA. The stimulation of each of these receptors leads to
activation of the transcription factor NF-.kappa.B, and other
signaling molecules that are involved in regulating the expression
of cytokine genes, including those encoding tumor necrosis
factor-alpha (TNF-.alpha.), interleukin-1 (IL-1), and certain
chemokines.
SUMMARY OF THE INVENTION
The present invention provides compounds and methods for use in
preventing or treating diseases or conditions characterized by
Toll-like receptor 2 (TLR2) activation in patients. Accordingly, in
a first aspect, the invention features a compound of formula I:
##STR00001## or a pharmaceutically acceptable salt or prodrug
thereof, where
a is an integer of 1 to 3;
b is an integer of 0 to 4, wherein when b is 0, the carbon bonded
to X and W is not bonded to 2 or more heteroatoms;
R.sup.1 is H or C.sub.1-6 alkyl;
X is selected from the group consisting of --NR.sup.X1V,
--N(R.sup.X1)C(O)V, --N(R.sup.X1)C(S)V,
--N(R.sup.X1)C(O)N(R.sup.X2)V, --N(R.sup.X1)C(S)N(R.sup.X2)V,
--N(R.sup.X1)C(O)OV, --N(R.sup.X1)S(O).sub.2V, --C(O)N(R.sup.X1)V,
--C(O)OV, --OC(O)V, --OC(O)OV, and --OC(O)N(R.sup.X1)V, where each
of R.sup.X1 and R.sup.X2 is, independently, H or C.sub.1-6 alkyl,
and V is a C.sub.1-20 alkyl, C.sub.1-20 alkenyl, or C.sub.1-20
alkynyl group, optionally substituted with halo, hydroxyl,
C.sub.1-21 acyloxy, oxo, C.sub.1-20 alkoxyl, or C.sub.1-20
thioalkoxyl, and optionally contains 1 to 2 phenyl or biphenyl
moieties in and/or at the end of the carbon chain;
W is selected from the group consisting of H,
--C(O)N(R.sup.W1)R.sup.W2, --C(O)OR.sup.W2,
--(CH.sub.2).sub.cOR.sup.W3, --(CH.sub.2).sub.cSR.sup.W3,
--(CH.sub.2).sub.cO(CH.sub.2).sub.dCH(OR.sup.W3)R.sup.W4,
--(CH.sub.2).sub.cS(CH.sub.2).sub.dCH(OR.sup.W3)R.sup.W4,
--C(O)N(R.sup.W1)(CH.sub.2).sub.cCH(OR.sup.W3)R.sup.W4, and
--C(O)N(R.sup.W1)(CH.sub.2).sub.cCH(OR.sup.W3)(CH.sub.2).sub.eOR.sup.W5,
where each of c and d is an integer of 1 to 4, e is an integer of 2
to 4, R.sup.W1 is H or C.sub.1-6 alkyl, R.sup.W2 is C.sub.1-20
alkyl, C.sub.1-20 alkenyl, or C.sub.1-20 alkynyl, each of R.sup.W3
and R.sup.W5 is, independently, H, C.sub.1-20 alkyl, C.sub.1-21
acyl, C.sub.1-20 alkenyl, or C.sub.1-20 alkynyl, and R.sup.W4 is H,
C.sub.1-20 alkyl, C.sub.1-20 alkenyl, or C.sub.1-20 alkynyl, where
each of R.sup.W2, R.sup.W3, R.sup.W4, and R.sup.W5 is optionally
substituted with halo, hydroxyl, C.sub.1-21 acyloxy, oxo,
C.sub.1-20 alkoxyl, or C.sub.1-20 thioalkoxyl, optionally contains
1 to 2 phenyl or biphenyl moieties in and/or at the end of the
carbon chain, and optionally contains 1 to 4 non-vicinal oxygen
atoms in the carbon chain; and
U is selected from the group consisting of
##STR00002## where
f is an integer of 1 to 4, g is an integer of 0 to 1,
each of R.sup.U1, R.sup.U2, and R.sup.U3 is, independently, H,
optionally substituted C.sub.1-6 alkyl, optionally substituted
C.sub.7-16 aralkyl, or optionally substituted C.sub.2-15
heterocyclylalkyl, or R.sup.U1 is H or optionally substituted
C.sub.1-6 alkyl and R.sup.U2 and R.sup.U3 together with the carbon
atom they are bonded to form an optionally substituted C.sub.3-6
aliphatic ring, or R.sup.U2 is H and R.sup.U3 and R.sup.U1 together
with the carbon atom bonded to R.sup.U3 and the nitrogen atom
bonded to R.sup.U1 form an optionally substituted 4-6-membered
heterocyclic ring,
R.sup.U4 is selected from the group consisting of
--CH.sub.2R.sup.U5, --C(O)R.sup.U6, --C(O)NH(R.sup.U7), and
--C(O)O(R.sup.U8), where each of R.sup.U5, R.sup.U6, R.sup.U7, and
R.sup.U8 is selected from the group consisting of optionally
substituted C.sub.1-6 alkyl, optionally substituted C.sub.2-6
alkenyl, optionally substituted C.sub.2-6 alkynyl, optionally
substituted C.sub.7-16 aralkyl, optionally substituted C.sub.2-15
heterocyclylalkyl, optionally substituted C.sub.6-10 aryl, and
optionally substituted C.sub.1-9 heterocyclyl, or R.sup.U4 is a
peptide chain of 1 to 10 natural or non-natural amino acids, or
mixture thereof, linked via the C-terminal end and substituted at
the N-terminal end of the peptide with a group selected from H,
--CH.sub.2R.sup.U5, --C(O)R.sup.U6, --C(O)NH(R.sup.U7), and
--C(O)O(R.sup.U8), where each of R.sup.U5, R.sup.U6, R.sup.U7, and
R.sup.U8 is as defined above, and
R.sup.U5 is a peptide chain of 1 to 10 natural or non-natural amino
acids, or mixture thereof, linked via the N-terminal end and the
C-terminal end is CO.sub.2R.sup.U9, or CONR.sup.U10R.sup.U11, where
each of R.sup.U9, R.sup.U10, and R.sup.U11 is selected from the
group consisting of H, optionally substituted C.sub.1-6 alkyl,
optionally substituted C.sub.2-6 alkenyl, optionally substituted
C.sub.2-6 alkynyl, optionally substituted C.sub.7-16 aralkyl,
optionally substituted C.sub.2-15 heterocyclylalkyl, optionally
substituted C.sub.6-10 aryl, and optionally substituted C.sub.1-9
heterocyclyl.
In a second aspect, the invention features a compound of formula
II:
##STR00003## or a pharmaceutically acceptable salt or prodrug
thereof, where each of a, b, U, X, and W is as defined above for
the compound of formula I; each of R.sup.1, R.sup.2, and R.sup.3
is, independently, H or C.sub.1-4 alkyl; R.sup.4 is H, optionally
substituted C.sub.1-6 alkyl, optionally substituted C.sub.7-16
aralkyl, or optionally substituted C.sub.2-15 heterocyclylalkyl;
and R.sup.5 is CO.sub.2H, SO.sub.3H, OP(O)(OH).sub.2, OSO.sub.3H,
or 5-tetrazolyl.
In an embodiment of either the first or second aspect of the
invention, X or W contains at least one linear alkyl moiety of 7 or
more carbons. Preferably, each of X and W contains at least one
linear alkyl moiety of 7 or more carbons.
Examples of compounds of the invention where W contains at least
one linear alkyl moiety of 7 or more carbons include those compound
in which W is selected from the group consisting of:
--C(O)NH(CH.sub.2).sub.2CH(OH)R.sup.W4, where R.sup.W4 is
C.sub.7-19 alkyl; --C(O)NH(CH.sub.2).sub.2CH.sub.2OR.sup.W3, where
R.sup.W3 is --C(O)(CH.sub.2).sub.aaCH.sub.3 and where aa is an
integer of 6 to 18; and
--C(O)NH(CH.sub.2).sub.2CH(OR.sup.W3)R.sup.W4, where R.sup.W3 is
--C(O)(CH.sub.2).sub.aaCH.sub.3 and R.sup.W4 is
CH.sub.2OC(O)(CH.sub.2).sub.bbCH.sub.3, where each of aa and bb is,
independently, an integer of 6 to 18.
In another embodiment of either the first or second aspect of the
invention, U is C(O)C(R.sup.U2)(R.sup.U3)NHR.sup.U4 or
--C(O)(CH.sub.2).sub.fNHR.sup.U4, where f is an integer of 1 to 4,
R.sup.U2 is an optionally substituted C.sub.1-6alkyl, R.sup.U3 is
H, and R.sup.U4 is an optionally substituted C.sub.6-10 aryl or an
optionally substituted C.sub.2-9 heterocyclyl. Examples include
those compounds in which R.sup.U4 is
##STR00004## where
R.sup.U12 is optionally substituted C.sub.6-10 aryl, optionally
substituted C.sub.6-10 aryloxy, optionally substituted C.sub.7-16
aralkyl, optionally substituted C.sub.7-16 aralkoxy, optionally
substituted C.sub.2-9 heterocyclyl, optionally substituted
C.sub.2-9 heterocyclyloxy, optionally substituted C.sub.3-15
heterocyclylalkyl, or optionally substituted C.sub.3-15
heterocyclylalkyloxy. Most preferably, R.sup.U4 is selected from
the group consisting of:
##STR00005##
Other compounds of the invention include those selected from the
group consisting of:
##STR00006## ##STR00007## ##STR00008## ##STR00009## ##STR00010##
##STR00011##
In a third aspect, the invention features a compound having the
formula:
##STR00012## or a pharmaceutically acceptable salt or prodrug
thereof, wherein
i is an integer of 1 to 4
R.sup.6 is H or C.sub.1-6 alkyl;
Z is selected from the group consisting of --NR.sup.Z1V,
--N(R.sup.Z1)C(O)V, --N(R.sup.Z1)C(S)V,
--N(R.sup.Z1)C(O)N(R.sup.Z2)V, --N(R.sup.Z1)C(S)N(R.sup.Z2)V,
--N(R.sup.Z1)C(O)OV, and --N(R.sup.Z1)S(O).sub.2V, where each of
R.sup.Z1 and R.sup.Z2 is, independently, H or C.sub.1-4 alkyl, and
V is a C.sub.1-20 alkyl, C.sub.1-20 alkenyl, or C.sub.1-20 alkynyl
group, optionally substituted with halo, hydroxyl, C.sub.1-21
acyloxy, oxo, C.sub.1-20 alkoxyl, or C.sub.1-20 thioalkoxyl and
optionally contains 1 to 2 phenyl or biphenyl moieties in and/or at
the end of the carbon chain;
R.sup.7 is C.sub.1-20 alkyl, C.sub.1-20 alkenyl, or C.sub.1-20
alkynyl, optionally substituted with halo, hydroxyl, C.sub.1-21
acyloxy, oxo, C.sub.1-20 alkoxyl, or C.sub.1-20 thioalkoxyl and
optionally contains 1 to 2 phenyl or biphenyl moieties in and/or at
the end of the carbon chain;
each of R.sup.8 and R.sup.9 is, independently H, optionally
substituted C.sub.1-6 alkyl, optionally substituted C.sub.7-16
aralkyl, or optionally substituted C.sub.2-15 heterocyclylalkyl, or
R.sup.8 and R.sup.9 together with the carbon atom they are bonded
to form an optionally substituted C.sub.3-4 aliphatic ring; and
T is OR.sup.T1, NR.sup.T2R.sup.T3, or a peptide chain of 1 to 10
natural or non-natural amino acids, or mixture thereof, linked via
the N-terminal end and the C-terminal end is CO.sub.2R.sup.T1, or
CONR.sup.T2R.sup.T3, wherein each of R.sup.T1, R.sup.T2, and
R.sup.T3 is selected from the group consisting of H, optionally
substituted C.sub.1-6 alkyl, optionally substituted C.sub.2-6
alkenyl, optionally substituted C.sub.2-6 alkynyl, optionally
substituted C.sub.7-16 aralkyl, optionally substituted C.sub.2-15
heterocyclylalkyl, optionally substituted C.sub.6-10 aryl, and
optionally substituted C.sub.1-9 heterocyclyl.
The invention also features a pharmaceutical composition that
includes any of the compounds of the invention and a
pharmaceutically acceptable excipient. The pharmaceutical
compositions of the inventions can be used to treat or prevent a
disease or condition characterized by Toll-like receptor 2
activation in a mammal, such as, for example, a human patient.
Accordingly, the invention features a method for treating or
preventing a disease in a mammal having, or predisposed to having,
a condition characterized by Toll-like receptor 2 activation that
includes administering a compound of formula I or formula II to the
mammal in an amount sufficient to treat or prevent the disease or
condition. The therapeutic methods of the invention can also
involve administration of one or more compounds that is selective
for TLR2 over, for example, TLR4, as well as methods involving
administration of TLR2/TLR4 dual antagonists.
Examples of diseases or conditions characterized by TLR2 activation
and that can be treated according to the invention include
inflammatory bowel disease, sepsis, periodontal disease, mucositis,
acne, cardiovascular disease, chronic obstructive pulmonary
disease, arthritis, cystic fibrosis, bacterial-induced infections,
viral-induced infections, mycoplasma-associated diseases,
post-herpetic neuralgia, ischemia/reperfusion injury, asthma,
stroke, brain injury, necrotizing enterocolitis, bed sores,
leprosy, atopic dermatitis, psoriasis, trauma, allergy,
neurodegenerative disease, amphotericin B-induced fever and
nephritis, coronary artery bypass grafting, and
atherosclerosis.
The invention also includes methods for identifying agents that
decrease or inhibit activation of Toll-like receptor 2. These
methods involve (i) contacting a cell expressing the receptor with
a candidate agent in the presence of an activator of the receptor
(in vitro or in vivo) and (ii) determining the effect of the agent
on activation of the receptor. Detection of a decrease in
activation of the receptor by the activator in the presence of the
agent indicates the identification of agent that can be used to
decrease or inhibit activation of the receptor. In these methods,
the effect of the agent on the activation of the receptor can be
determined by analysis of the expression of a reporter gene that is
under the control of a promoter that is induced in a signaling
pathway triggered by activation of the receptor.
The terms "acyl" or "alkanoyl," as used interchangeably herein,
represent an alkyl group, as defined herein, or hydrogen attached
to the parent molecular group through a carbonyl group, as defined
herein, and is exemplified by formyl, acetyl, propionyl, butanoyl
and the like. Exemplary unsubstituted acyl groups are of 2 to 21
carbons.
The term "acyloxy" represents an alkyl group, as defined herein,
attached to the parent molecular group through a carbonyl group and
an oxygen atom. Exemplary acyloxy groups are of 2 to 21
carbons.
The term "alkenyl," as used herein, represents monovalent straight
or branched chain groups of, unless otherwise specified, from 2 to
20 carbons containing one or more carbon-carbon double bonds and is
exemplified by ethenyl, 1-propenyl, 2-propenyl,
2-methyl-1-propenyl, 1-butenyl, 2-butenyl and the like and may be
optionally substituted with one, two, three or four substituents
independently selected from the group consisting of: (1) alkoxy of
one to twenty carbon atoms; (2) alkylsulfinyl of one to twenty
carbon atoms; (3) alkylsulfonyl of one to twenty carbon atoms; (4)
amino; (5) aryl; (6) arylalkoxy, where the alkylene group is of one
to twenty carbon atoms; (7) aryloyl; (8) azido; (9) carboxaldehyde;
(10) cycloalkyl of three to eight carbon atoms; (11) halo; (12)
heterocycle; (13) (heterocycle)oxy; (14) (heterocycle)oyl; (15)
hydroxy; (16) N-protected amino; (17) nitro; (18) oxo; (19)
spiroalkyl of three to eight carbon atoms; (20) thioalkoxy of one
to twenty carbon atoms; (21) thiol; (22) --CO.sub.2R.sup.A, where
R.sup.A is selected from the group consisting of (a) hydrogen, (b)
substituted or unsubstituted C.sub.1-20 alkyl, (c) substituted or
unsubstituted C.sub.6 or C.sub.10 aryl, (d) substituted or
unsubstituted C.sub.7-16 arylalkyl, where the alkylene group is of
one to twenty carbon atoms, (e) substituted or unsubstituted
C.sub.1-9 heterocyclyl, and (f) substituted or unsubstituted
C.sub.2-15 heterocyclylalkyl, where the alkylene group is of one to
twenty carbon atoms; (23) --C(O)NR.sup.BR.sup.C, where each of
R.sup.B and R.sup.C is, independently, selected from the group
consisting of (a) hydrogen, (b) alkyl, (c) aryl, and (d) arylalkyl,
where the alkylene group is of one to twenty carbon atoms; (24)
--S(O).sub.2R.sup.D, where R.sup.D is selected from the group
consisting of (a) alkyl, (b) aryl, and (c) arylalkyl, where the
alkylene group is of one to twenty carbon atoms; (25)
--S(O).sub.2NR.sup.ER.sup.F, where each of R.sup.E and R.sup.F is,
independently, selected from the group consisting of (a) hydrogen,
(b) alkyl, (c) aryl, and (d) arylalkyl, where the alkylene group is
of one to twenty carbon atoms; and (26) --NR.sup.GR.sup.H, where
each of R.sup.G and R.sup.H is, independently, selected from the
group consisting of (a) hydrogen, (b) an N-protecting group, (c)
alkyl of one to twenty carbon atoms, (d) alkenyl of two to twenty
carbon atoms, (e) alkynyl of two to twenty carbon atoms, (f) aryl,
(g) arylalkyl, where the alkylene group is of one to twenty carbon
atoms, (h) cycloalkyl of three to eight carbon atoms, and (i)
cycloalkylalkyl, where the cycloalkyl group is of three to eight
carbon atoms, and the alkylene group is of one to ten carbon atoms,
with the proviso that no two groups are bound to the nitrogen atom
through a carbonyl group or a sulfonyl group.
The terms "alkoxy" or "alkyloxy," as used interchangeably herein,
represent an alkyl group attached to the parent molecular group
through an oxygen atom. Exemplary unsubstituted alkoxy groups are
of 1 to 20 carbons.
The term "alkyl," as used herein, represents a monovalent group
derived from a straight or branched chain saturated hydrocarbon of,
unless otherwise specified, from 1 to 20 carbons and is exemplified
by methyl, ethyl, n- and iso-propyl, n-, see-, iso- and tert-butyl,
neopentyl and the like and may be optionally substituted with one,
two, three or, in the case of alkyl groups of two carbons or more,
four substituents independently selected from the group consisting
of: (1) alkoxy of one to twenty carbon atoms; (2) alkylsulfinyl of
one to twenty carbon atoms; (3) alkylsulfonyl of one to twenty
carbon atoms; (4) amino; (5) aryl; (6) arylalkoxy; (7) aryloyl; (8)
azido; (9) carboxaldehyde; (10) cycloalkyl of three to eight carbon
atoms; (11) halo; (12) heterocyclyl; (13) (heterocycle)oxy; (14)
(heterocycle)oyl; (15) hydroxyl; (16) N-protected amino; (17)
nitro; (18) oxo; (19) spiroalkyl of three to eight carbon atoms;
(20) thioalkoxy of one to twenty carbon atoms; (21) thiol; (22)
--CO.sub.2R.sup.A, where R.sup.A is selected from the group
consisting of (a) hydrogen, (b) substituted or unsubstituted
C.sub.1-20 alkyl, (c) substituted or unsubstituted C.sub.6 or
C.sub.10 aryl, (d) substituted or unsubstituted C.sub.7-16
arylalkyl, where the alkylene group is of one to twenty carbon
atoms, (e) substituted or unsubstituted C.sub.1-9 heterocyclyl, and
(f) substituted or unsubstituted C.sub.2-15 heterocyclylalkyl,
where the alkylene group is of one to twenty carbon atoms; (23)
--C(O)NR.sup.BR.sup.C, where each of R.sup.B and R.sup.C is,
independently, selected from the group consisting of (a) hydrogen,
(b) alkyl, (c) aryl, and (d) arylalkyl, where the alkylene group is
of one to twenty carbon atoms; (24) --S(O).sub.2R.sup.D, where
R.sup.D is selected from the group consisting of (a) alkyl, (b)
aryl, and (c) arylalkyl, where the alkylene group is of one to
twenty carbon atoms; (25) --S(O).sub.2NR.sup.ER.sup.F, where each
of R.sup.E and R.sup.F is, independently, selected from the group
consisting of (a) hydrogen, (b) alkyl, (c) aryl, and (d) arylalkyl,
where the alkylene group is of one to twenty carbon atoms; and (26)
--NR.sup.GR.sup.H, where each of R.sup.G and R.sup.H is,
independently, selected from the group consisting of (a) hydrogen,
(b) an N-protecting group, (c) alkyl of one to twenty carbon atoms,
(d) alkenyl of two to twenty carbon atoms, (e) alkynyl of two to
twenty carbon atoms, (f) aryl, (g) arylalkyl, where the alkylene
group is of one to twenty carbon atoms, (h) cycloalkyl of three to
eight carbon atoms, and (i) cycloalkylalkyl, where the cycloalkyl
group is of three to eight carbon atoms, and the alkylene group is
of one to ten carbon atoms, with the proviso that no two groups are
bound to the nitrogen atom through a carbonyl group or a sulfonyl
group.
The term "alkylene," as used herein, represents a saturated
divalent hydrocarbon group derived from a straight or branched
chain saturated hydrocarbon by the removal of two hydrogen atoms,
and is exemplified by methylene, ethylene, isopropylene and the
like.
The term "alkylthio," as used herein, represents an alkyl group
attached to the parent molecular group through a sulfur atom.
Exemplary unsubstituted alkylthio groups are of 1 to 20
carbons.
The term "alkynyl," as used herein, represents monovalent straight
or branched chain groups of 2 to 20 carbon atoms containing a
carbon-carbon triple bond and is exemplified by ethynyl,
1-propynyl, and the like and may be optionally substituted with
one, two, three or four substituents independently selected from
the group consisting of: (1) alkoxy of one to twenty carbon atoms;
(2) alkylsulfinyl of one to twenty carbon atoms; (3) alkylsulfonyl
of one to twenty carbon atoms; (4) amino; (5) aryl; (6) arylalkoxy,
where the alkylene group is of one to twenty carbon atoms; (7)
aryloyl; (8) azido; (9) carboxaldehyde; (10) cycloalkyl of three to
eight carbon atoms; (11) halo; (12) heterocycle; (13)
(heterocycle)oxy; (14) (heterocycle)oyl; (15) hydroxy; (16)
N-protected amino; (17) nitro; (18) oxo; (19) spiroalkyl of three
to eight carbon atoms; (20) thioalkoxy of one to twenty carbon
atoms; (21) thiol; (22) --CO.sub.2R.sup.A, where R.sup.A is
selected from the group consisting of (a) alkyl, (b) aryl and (c)
arylalkyl, where the alkylene group is of one to twenty carbon
atoms; (23) --C(O)NR.sup.BR.sup.C, where each of R.sup.B and
R.sup.C is, independently, selected from the group consisting of
(a) hydrogen, (b) alkyl, (c) aryl, and (d) arylalkyl, where the
alkylene group is of one to twenty carbon atoms; (24)
--S(O).sub.2R.sup.D, where R.sup.D is selected from the group
consisting of (a) alkyl, (b) aryl, and (c) arylalkyl, where the
alkylene group is of one to twenty carbon atoms; (25)
--S(O).sub.2NR.sup.ER.sup.F, where each of R.sup.E and R.sup.F is,
independently, selected from the group consisting of (a) hydrogen,
(b) alkyl, (c) aryl, and (d) arylalkyl, where the alkylene group is
of one to twenty carbon atoms; and (26) --NR.sup.GR.sup.H, where
each of R.sup.G and R.sup.H is, independently, selected from the
group consisting of (a) hydrogen, (b) an N-protecting group, (c)
alkyl of one to twenty carbon atoms, (d) alkenyl of two to twenty
carbon atoms, (e) alkynyl of two to twenty carbon atoms, (f) aryl,
(g) arylalkyl, where the alkylene group is of one to twenty carbon
atoms, (h) cycloalkyl of three to eight carbon atoms, and (i)
cycloalkylalkyl, where the cycloalkyl group is of three to eight
carbon atoms, and the alkylene group is of one to ten carbon atoms,
with the proviso that no two groups are bound to the nitrogen atom
through a carbonyl group or a sulfonyl group.
The term "alpha-amino acid residue," as used herein, represents a
--N(R.sup.A)C(R.sup.B)(R.sup.C)C(O)-- linkage, where R.sup.A is
selected from the group consisting of (a) hydrogen, (b) alkyl, (c)
aryl, and (d) arylalkyl, as defined herein; and each of R.sup.B and
R.sup.C is, independently, selected from the group consisting of:
(a) hydrogen, (b) optionally substituted alkyl, (c) optionally
substituted cycloalkyl, (d) optionally substituted aryl, (e)
optionally substituted arylalkyl, (f) optionally substituted
heterocyclyl, and (g) optionally substituted heterocyclylalkyl,
each of which is as defined herein. For natural amino acids,
R.sup.B is H and R.sup.C corresponds to those side chains of
natural amino acids found in nature, or their antipodal
configurations. Exemplary natural amino acids include alanine,
cysteine, aspartic acid, glutamic acid, phenylalanine, glycine,
histidine, isoleucine, lysine, leucine, methionine, aspartamine,
ornithine, proline, glutamine, arginine, serine, threonine, valine,
tryptophan, and tyrosine, each of which, except glycine, as their
D- or L-form. As used herein, for the most part, the names of
naturally-occurring amino acids and aminoacyl residues used herein
follow the naming conventions suggested by the IUPAC Commission on
the Nomenclature of Organic Chemistry and the IUPAC-IUB Commission
on Biochemical Nomenclature as set out in Nomenclature of
.alpha.-Amino Acids (Recommendations, 1974), Biochemistry 14 (2),
1975. The present invention also contemplates non-naturally
occurring (i.e., non-natural) amino acid residues in their D- or
L-form such as, for example, homophenylalanine, phenylglycine,
cyclohexylglycine, cyclohexylalanine, cyclopentyl alanine,
cyclobutylalanine, cyclopropylalanine, cyclohexylglycine,
norvaline, norleucine, thiazoylalanine (2-, 4-, and 5-substituted),
pyridylalanine (2-, 3-, and 4-isomers), naphthalalanine (1- and
2-isomers) and the like. Non-natural amino acids also include
beta-amino acids, optionally substituted at the alpha or beta or
both alpha and beta positions, independently, with R.sup.A and
R.sup.B, as described above.
The term "amino," as used herein, represents an --NH.sub.2
group.
The term "aryl," as used herein, represents a mono- or bicyclic
carbocyclic ring system having one or two aromatic rings and is
exemplified by phenyl, naphthyl, 1,2-dihydronaphthyl,
1,2,3,4-tetrahydronaphthyl, fluorenyl, indanyl, indenyl and the
like and may be optionally substituted with one, two, three, four,
or five substituents independently selected from the group
consisting of: (1) alkanoyl of one to twenty carbon atoms; (2)
alkyl of one to twenty carbon atoms; (3) alkoxy of one to twenty
carbon atoms; (4) alkoxyalkyl, where the alkyl and alkylene groups
are independently of one to twenty carbon atoms; (5) alkylsulfinyl
of one to twenty carbon atoms; (6) alkylsulfinylalkyl, where the
alkyl and alkylene groups are independently of one to twenty carbon
atoms; (7) alkylsulfonyl of one to twenty carbon atoms; (8)
alkylsulfonylalkyl, where the alkyl and alkylene groups are
independently of one to twenty carbon atoms; (9) aryl; (10)
arylalkyl, where the alkyl group is of one to twenty carbon atoms;
(11) amino; (12) aminoalkyl of one to twenty carbon atoms; (13)
aryl; (14) arylalkyl, where the alkylene group is of one to twenty
carbon atoms; (15) aryloyl; (16) azido; (17) azidoalkyl of one to
twenty carbon atoms; (18) carboxaldehyde; (19)
(carboxaldehyde)alkyl, where the alkylene group is of one to twenty
carbon atoms; (20) cycloalkyl of three to eight carbon atoms; (21)
cycloalkylalkyl, where the cycloalkyl group is of three to eight
carbon atoms and the alkylene group is of one to ten carbon atoms;
(22) halo; (23) haloalkyl of one to twenty carbon atoms; (24)
heterocyclyl; (25) (heterocyclyl)oxy; (26) (heterocyclyl)oyl; (27)
hydroxy; (28) hydroxyalkyl of one to twenty carbon atoms; (29)
nitro; (30) nitroalkyl of one to twenty carbon atoms; (31)
N-protected amino; (32) N-protected aminoalkyl, where the alkylene
group is of one to twenty carbon atoms; (33) oxo; (34) thioalkoxy
of one to twenty carbon atoms; (35) thioalkoxyalkyl, where the
alkyl and alkylene groups are independently of one to twenty carbon
atoms; (36) --(CH.sub.2).sub.qCO.sub.2R.sup.A, where q is an
integer of zero to four and R.sup.A is selected from the group
consisting of (a) alkyl, (b) aryl, and (c) arylalkyl, where the
alkylene group is of one to twenty carbon atoms; (37)
--(CH.sub.2).sub.qCONR.sup.BR.sup.C, where R.sup.B and R.sup.C are
independently selected from the group consisting of (a) hydrogen,
(b) alkyl, (c) aryl, and (d) arylalkyl, where the alkylene group is
of one to twenty carbon atoms; (38)
--(CH.sub.2).sub.qS(O).sub.2R.sup.D, where R.sup.D is selected from
the group consisting of (a) alkyl, (b) aryl, and (c) arylalkyl,
where the alkylene group is of one to twenty carbon atoms; (39)
--(CH.sub.2).sub.qS(O).sub.2NR.sup.ER.sup.F, where each of R.sup.E
and R.sup.F is, independently, selected from the group consisting
of (a) hydrogen, (b) alkyl, (c) aryl, and (d) arylalkyl, where the
alkylene group is of one to twenty carbon atoms; (40)
--(CH.sub.2).sub.qNR.sup.GR.sup.H, where each of R.sup.G and
R.sup.H is, independently, selected from the group consisting of
(a) hydrogen, (b) an N-protecting group, (c) alkyl of one to twenty
carbon atoms, (d) alkenyl of two to twenty carbon atoms, (e)
alkynyl of two to twenty carbon atoms, (f) aryl, (g) arylalkyl,
where the alkylene group is of one to twenty carbon atoms, (h)
cycloalkyl of three to eight carbon atoms, and (i) cycloalkylalkyl,
where the cycloalkyl group is of three to eight carbon atoms, and
the alkylene group is of one to ten carbon atoms, with the proviso
that no two groups are bound to the nitrogen atom through a
carbonyl group or a sulfonyl group; (41) oxo; (42) thiol; (43)
perfluoroalkyl; (44) perfluoroalkoxy; (45) aryloxy; (46)
cycloalkoxy; (47) cycloalkylalkoxy; and (48) arylalkoxy.
The terms "arylalkyl" or "aralkyl," as used interchangeably herein,
represent an aryl group attached to the parent molecular group
through an alkyl group. Exemplary unsubstituted arylalkyl groups
are of 7 to 16 carbons.
The term "aryloxy," as used herein, represents an aryl group that
is attached to the parent molecular group through an oxygen atom.
Exemplary unsubstituted aryloxy groups are of 6 or 10 carbons.
The terms "aryloyl" or "aroyl," as used interchangeably herein,
represent an aryl group that is attached to the parent molecular
group through a carbonyl group. Exemplary unsubstituted
aryloxycarbonyl groups are of 7 or 11 carbons.
The term "carbonyl" as used herein, represents a C.dbd.O group.
The term "carboxy" or "carboxyl," as used interchangeably herein,
represents a --CO.sub.2H group.
The terms "carboxy protecting group" or "carboxyl protecting
group," as used herein, represent those groups intended to protect
a --CO.sub.2H group against undesirable reactions during synthetic
procedures. Commonly used carboxy-protecting groups are disclosed
in Greene, "Protective Groups In Organic Synthesis, 3rd Edition"
(John Wiley & Sons, New York, 1999), which is incorporated
herein by reference.
The phrase "compound selective for antagonism of Toll-like receptor
2 over Toll-like receptor 4" is used to describe those compounds
that have an IC.sub.50 value when tested by the TLR2 in vitro assay
described herein that is less than the IC.sub.50 value obtained
when the compound is tested by the TLR4 in vitro assay described
herein.
The term "cycloalkyl," as used herein, represents a monovalent
saturated or unsaturated non-aromatic cyclic hydrocarbon group of
three to eight carbons, unless otherwise specified, and is
exemplified by cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl,
cycloheptyl, bicyclo[2.2.1.]heptyl and the like. The cycloalkyl
groups of this invention can be optionally substituted with (1)
alkanoyl of one to twenty carbon atoms; (2) alkyl of one to twenty
carbon atoms; (3) alkoxy of one to twenty carbon atoms; (4)
alkoxyalkyl, where the alkyl and alkylene groups are independently
of one to twenty carbon atoms; (5) alkylsulfinyl of one to twenty
carbon atoms; (6) alkylsulfinylalkyl, where the alkyl and alkylene
groups are independently of one to twenty carbon atoms; (7)
alkylsulfonyl of one to twenty carbon atoms; (8)
alkylsulfonylalkyl, where the alkyl and alkylene groups are
independently of one to twenty carbon atoms; (9) aryl; (10)
arylalkyl, where the alkyl group is of one to twenty carbon atoms;
(11) amino; (12) aminoalkyl of one to twenty carbon atoms; (13)
aryl; (14) arylalkyl, where the alkylene group is of one to twenty
carbon atoms; (15) aryloyl; (16) azido; (17) azidoalkyl of one to
twenty carbon atoms; (18) carboxaldehyde; (19)
(carboxaldehyde)alkyl, where the alkylene group is of one to twenty
carbon atoms; (20) cycloalkyl of three to eight carbon atoms; (21)
cycloalkylalkyl, where the cycloalkyl group is of three to eight
carbon atoms and the alkylene group is of one to ten carbon atoms;
(22) halo; (23) haloalkyl of one to twenty carbon atoms; (24)
heterocyclyl; (25) (heterocyclyl)oxy; (26) (heterocyclyl)oyl; (27)
hydroxy; (28) hydroxyalkyl of one to twenty carbon atoms; (29)
nitro; (30) nitroalkyl of one to twenty carbon atoms; (31)
N-protected amino; (32) N-protected aminoalkyl, where the alkylene
group is of one to twenty carbon atoms; (33) oxo; (34) thioalkoxy
of one to twenty carbon atoms; (35) thioalkoxyalkyl, where the
alkyl and alkylene groups are independently of one to twenty carbon
atoms; (36) --(CH.sub.2).sub.qCO.sub.2R.sup.A, where q is an
integer of zero to four and R.sup.A is selected from the group
consisting of (a) alkyl, (b) aryl, and (c) arylalkyl, where the
alkylene group is of one to twenty carbon atoms; (37)
--(CH.sub.2).sub.qCONR.sup.BR.sup.C, where each of R.sup.B and
R.sup.C is, independently, selected from the group consisting of
(a) hydrogen, (b) alkyl, (c) aryl, and (d) arylalkyl, where the
alkylene group is of one to twenty carbon atoms; (38)
--(CH.sub.2).sub.qS(O).sub.2R.sup.D, where R.sup.D is selected from
the group consisting of (a) alkyl, (b) aryl, and (c) arylalkyl,
where the alkylene group is of one to twenty carbon atoms; (39)
--(CH.sub.2).sub.qS(O).sub.2NR.sup.ER.sup.F, where each of R.sup.E
and R.sup.F is, independently, selected from the group consisting
of (a) hydrogen, (b) alkyl, (c) aryl, and (d) arylalkyl, where the
alkylene group is of one to twenty carbon atoms; (40)
--(CH.sub.2).sub.qNR.sup.GR.sup.H, where each of R.sup.G and
R.sup.H is, independently, selected from the group consisting of
(a) hydrogen; (b) an N-protecting group; (c) alkyl of one to twenty
carbon atoms; (d) alkenyl of two to twenty carbon atoms; (e)
alkynyl of two to twenty carbon atoms; (f) aryl; (g) arylalkyl,
where the alkylene group is of one to twenty carbon atoms; (h)
cycloalkyl of three to eight carbon atoms and (i) cycloalkylalkyl,
where the cycloalkyl group is of three to eight carbon atoms, and
the alkylene group is of one to ten carbon atoms, with the proviso
that no two groups are bound to the nitrogen atom through a
carbonyl group or a sulfonyl group; (41) oxo; (42) thiol; (43)
perfluoroalkyl; (44) perfluoroalkoxy; (45) aryloxy; (46)
cycloalkoxy; (47) cycloalkylalkoxy; and (48) arylalkoxy.
The term "halogen" or "halo," as used interchangeably herein,
represents F, Cl, Br, and I.
The term "heteroaryl," as used herein, represents that subset of
heterocycles, as defined herein, which are aromatic: i.e., they
contain 4n+2 pi electrons within the mono- or multicyclic ring
system. Exemplary unsubstituted heteroaryl groups are of 1 to 9
carbons.
The terms "heterocycle" or "heterocyclyl," as used interchangeably
herein, represent a 5-, 6- or 7-membered ring, unless otherwise
specified, containing one, two, three, or four heteroatoms
independently selected from the group consisting of nitrogen,
oxygen and sulfur. The 5-membered ring has zero to two double bonds
and the 6- and 7-membered rings have zero to three double bonds.
The term "heterocycle" also includes bicyclic, tricyclic, and
tetracyclic groups in which any of the above heterocyclic rings is
fused to one or two rings independently selected from the group
consisting of an aryl ring, a cyclohexane ring, a cyclohexene ring,
a cyclopentane ring, a cyclopentene ring, and another monocyclic
heterocyclic ring such as indolyl, quinolyl, isoquinolyl,
tetrahydroquinolyl, benzofuryl, benzothienyl and the like.
Heterocyclics include pyrrolyl, pyrrolinyl, pyrrolidinyl,
pyrazolyl, pyrazolinyl, pyrazolidinyl, imidazolyl, imidazolinyl,
imidazolidinyl, pyridyl, piperidinyl, homopiperidinyl, pyrazinyl,
piperazinyl, pyrimidinyl, pyridazinyl, oxazolyl, oxazolidinyl,
isoxazolyl, isoxazolidiniyl, morpholinyl, thiomorpholinyl,
thiazolyl, thiazolidinyl, isothiazolyl, isothiazolidinyl, indolyl,
quinolinyl, isoquinolinyl, benzimidazolyl, benzothiazolyl,
benzoxazolyl, furyl, thienyl, thiazolidinyl, isothiazolyl,
isoindazoyl, triazolyl, tetrazolyl, oxadiazolyl, uricyl,
thiadiazolyl, pyrimidyl, tetrahydrofuranyl, dihydrofuranyl,
tetrahydrothienyl, dihydrothienyl, dihydroinidolyl,
tetrahydroquinolyl, tetrahydroisoquinolyl, pyranyl, dihydropyranyl,
dithiazolyl, benzofuranyl, benzothienyl, and the like. Heterocyclic
groups also include compounds of the formula
##STR00013##
F' is selected from the group consisting of --CH.sub.2--,
--CH.sub.2O--, and --O--, and G' is selected from the group
consisting of --C(O)-- and --(C(R')(R'')).sub.v--, where each of R'
and R'' is, independently, selected from the group consisting of
hydrogen or alkyl of one to four carbon atoms, and v is one to
three and includes groups such as 1,3-benzodioxolyl,
1,4-benzodioxanyl and the like. Any of the heterocycle groups
mentioned herein may be optionally substituted with one, two,
three, four, or five substituents independently selected from the
group consisting of: (1) alkanoyl of one to twenty carbon atoms;
(2) alkyl of one to twenty carbon atoms; (3) alkoxy of one to
twenty carbon atoms; (4) alkoxyalkyl, where the alkyl and alkylene
groups are independently of one to twenty carbon atoms; (5)
alkylsulfinyl of one to twenty carbon atoms; (6)
alkylsulfinylalkyl, where the alkyl and alkylene groups are
independently of one to twenty carbon atoms; (7) alkylsulfonyl of
one to twenty carbon atoms; (8) alkylsulfonylalkyl, where the alkyl
and alkylene groups are independently of one to twenty carbon
atoms; (9) aryl; (10) arylalkyl, where the alkyl group is of one to
twenty carbon atoms; (11) amino; (12) aminoalkyl of one to twenty
carbon atoms; (13) aryl; (14) arylalkyl, where the alkylene group
is of one to twenty carbon atoms; (15) aryloyl; (16) azido; (17)
azidoalkyl of one to twenty carbon atoms; (18) carboxaldehyde; (19)
(carboxaldehyde)alkyl, where the alkylene group is of one to twenty
carbon atoms; (20) cycloalkyl of three to eight carbon atoms; (21)
cycloalkylalkyl, where the cycloalkyl group is of three to eight
carbon atoms and the alkylene group is of one to ten carbon atoms;
(22) halo; (23) haloalkyl of one to twenty carbon atoms; (24)
heterocycle; (25) (heterocycle)oxy; (26) (heterocycle)oyl; (27)
hydroxy; (28) hydroxyalkyl of one to twenty carbon atoms; (29)
nitro; (30) nitroalkyl of one to twenty carbon atoms; (31)
N-protected amino; (32) N-protected aminoalkyl, where the alkylene
group is of one to twenty carbon atoms; (33) oxo; (34) thioalkoxy
of one to twenty carbon atoms; (35) thioalkoxyalkyl, where the
alkyl and alkylene groups are independently of one to twenty carbon
atoms; (36) --(CH.sub.2).sub.qCO.sub.2R.sup.A, where q is an
integer of zero to four and R.sup.A is selected from the group
consisting of (a) alkyl, (b) aryl, and (c) arylalkyl, where the
alkylene group is of one to twenty carbon atoms; (37)
--(CH.sub.2).sub.qCONR.sup.BR.sup.C, where each of R.sup.B and
R.sup.C is, independently, selected from the group consisting of
(a) hydrogen, (b) alkyl, (c) aryl, and (d) arylalkyl, where the
alkylene group is of one to twenty carbon atoms; (38)
--(CH.sub.2).sub.qS(O).sub.2R.sup.D, where R.sup.D is selected from
the group consisting of (a) alkyl, (b) aryl, and (c) arylalkyl,
where the alkylene group is of one to twenty carbon atoms; (39)
--(CH.sub.2).sub.qS(O).sub.2NR.sup.ER.sup.F, where each of R.sup.E
and R.sup.F is, independently, selected from the group consisting
of (a) hydrogen, (b) alkyl, (c) aryl, and (d) arylalkyl, where the
alkylene group is of one to twenty carbon atoms; (40)
--(CH.sub.2).sub.qNR.sup.GR.sup.H, where each of R.sup.G and
R.sup.H is, independently, selected from the group consisting of
(a) hydrogen; (b) an N-protecting group; (c) alkyl of one to twenty
carbon atoms; (d) alkenyl of two to twenty carbon atoms; (e)
alkynyl of two to twenty carbon atoms; (f) aryl; (g) arylalkyl,
where the alkylene group is of one to twenty carbon atoms; (h)
cycloalkyl of three to eight carbon atoms and (i) cycloalkylalkyl,
where the cycloalkyl group is of three to eight carbon atoms, and
the alkylene group is of one to ten carbon atoms, with the proviso
that no two groups are bound to the nitrogen atom through a
carbonyl group or a sulfonyl group; (41) oxo; (42) thiol; (43)
perfluoroalkyl; (44) perfluoroalkoxy; (45) aryloxy; (46)
cycloalkoxy; (47) cycloalkylalkoxy; and (48) arylalkoxy.
The term "heterocyclyloxy," as used herein, represents a
hetercyclyl group which is attached to the parent molecular group
through an oxygen atom.
The term "hydroxy" or "hydroxyl," as used interchangeably herein,
represents an --OH group.
The term "N-protected amino," as used herein, refers to an amino
group, as defined herein, to which is attached an N-protecting or
nitrogen-protecting group, as defined herein.
The terms "N-protecting group," "nitrogen protecting group," or
"amino protecting group," as used herein, represent those groups
intended to protect an amino group against undesirable reactions
during synthetic procedures. Commonly used N-protecting groups are
disclosed in Greene, "Protective Groups In Organic Synthesis, 3d
Edition" (John Wiley & Sons, New York, 1999), which is
incorporated herein by reference. N-protecting groups comprise
acyl, aroyl, or carbamyl groups such as formyl, acetyl, propionyl,
pivaloyl, t-butylacetyl, 2-chloroacetyl, 2-bromoacetyl,
trifluoroacetyl, trichloroacetyl, phthalyl, o-nitrophenoxyacetyl,
.alpha.-chlorobutyryl, benzoyl, 4-chlorobenzoyl, 4-bromobenzoyl,
4-nitrobenzoyl, and chiral auxiliaries such as protected or
unprotected L- or D-amino acids such as alanine, leucine,
phenylalanine, and the like; sulfonyl groups such as
benzenesulfonyl, p-toluenesulfonyl and the like; carbamate forming
groups such as benzyloxycarbonyl, p-chlorobenzyloxycarbonyl,
p-methoxybenzyloxycarbonyl, p-nitrobenzyloxycarbonyl,
2-nitrobenzyloxycarbonyl, p-bromobenzyloxycarbonyl,
3,4-dimethoxybenzyloxycarbonyl, 3,5-dimethoxybenzyloxycarbonyl,
2,4-dimethoxybenzyloxycarbonyl, 4-methoxybenzyloxycarbonyl,
2-nitro-4,5-dimethoxybenzyloxycarbonyl,
3,4,5-trimethoxybenzyloxycarbonyl,
1-(p-biphenylyl)-1-methylethoxycarbonyl,
.alpha.,.alpha.-dimethyl-3,5-dimethoxybenzyloxycarbonyl,
benzhydryloxycarbonyl, t-butyloxycarbonyl,
diisopropylmethoxycarbonyl, isopropyloxycarbonyl, ethoxycarbonyl,
methoxycarbonyl, allyloxycarbonyl, 2,2,2-trichloroethoxycarbonyl,
phenoxycarbonyl, 4-nitrophenoxy carbonyl,
fluorenyl-9-methoxycarbonyl, cyclopentyloxycarbonyl,
adamantyloxycarbonyl, cyclohexyloxycarbonyl, phenylthiocarbonyl,
and the like, arylalkyl groups such as benzyl, triphenylmethyl,
benzyloxymethyl, and the like and silyl groups such as
trimethylsilyl and the like. Preferred N-protecting groups are
formyl, acetyl, benzoyl, pivaloyl, t-butylacetyl, alanyl,
phenylsulfonyl, benzyl, t-butyloxycarbonyl (Boc), and
benzyloxycarbonyl (Cbz).
The term "non-vicinal oxygen atoms" refers to oxygen atoms that are
not bonded to the same carbon atom.
The term "pharmaceutically acceptable salt," as use herein, refers
to those salts which are, within the scope of sound medical
judgement, suitable for use in contact with the tissues of humans
and animals without undue toxicity, irritation, allergic response
and the like and are commensurate with a reasonable benefit/risk
ratio. Pharmaceutically acceptable salts are well known in the art.
For example, S. M. Berge et al. describe pharmaceutically
acceptable salts in detail in J. Pharmaceutical Sciences 66:1-19,
1977. The salts can be prepared in situ during the final isolation
and purification of a compound of the invention or separately by
reacting the free base group with a suitable organic acid.
Representative acid addition salts include acetate, adipate,
alginate, ascorbate, aspartate, benzenesulfonate, benzoate,
bisulfate, borate, butyrate, camphorate, camphersulfonate, citrate,
cyclopentanepropionate, digluconate, dodecylsulfate,
ethanesulfonate, fumarate, glucoheptonate, glycerophosphate,
hemisulfate, heptonate, hexanoate, hydrobromide, hydrochloride,
hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate, lactate,
laurate, lauryl sulfate, malate, maleate, malonate,
methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate,
oleate, oxalate, palmitate, pamoate, pectinate, persulfate,
3-phenylpropionate, phosphate, picrate, pivalate, propionate,
stearate, succinate, sulfate, tartrate, thiocyanate,
toluenesulfonate, undecanoate, valerate salts, and the like.
Representative alkali or alkaline earth metal salts include sodium,
lithium, potassium, calcium, magnesium and the like, as well as
nontoxic ammonium, quaternary ammonium, and amine cations,
including, but not limited to ammonium, tetramethylammonium,
tetraethylammonium, methylamine, dimethylamine, trimethylamine,
triethylamine, ethylamine, and the like. The term "pharmaceutically
acceptable ester," as used herein, represents esters that hydrolyze
in vivo and include those that break down readily in the human body
to leave the parent compound or a salt thereof. Suitable ester
groups include, for example, those derived from pharmaceutically
acceptable aliphatic carboxylic acids, particularly alkanoic,
alkenoic, cycloalkanoic, and alkanedioic acids, in which each alkyl
or alkenyl group preferably has not more than 6 carbon atoms.
Examples of particular esters include formates, acetates,
propionates, butyates, acrylates, and ethylsuccinates.
The term "pharmaceutically acceptable prodrugs," as used herein,
means prodrugs of the compounds of the present invention which are,
within the scope of sound medical judgement, suitable for use in
contact with the tissues of humans and animals with undue toxicity,
irritation, allergic response, and the like, commensurate with a
reasonable benefit/risk ratio, and effective for their intended
use, as well as the zwitterionic forms, where possible, of the
compounds of the invention.
The term "phenyl" means an aromatic ring containing 6 carbons.
Phenyl rings can be optionally substituted. When a phenyl ring is
in a carbon chain it is part of the carbon chain linkage (i.e., the
phenyl ring is bonded to the chain at two positions in either an
ortho, meta, or para fashion). When a phenyl ring is at the end of
a carbon chain, it is bonded to the end of the carbon chain.
The term "prodrug," as used herein, represents compounds that are
transformed in vivo into a parent compound of the above formula,
for example, by hydrolysis in blood. A thorough discussion of
prodrugs is provided in T. Higuchi and V. Stella, "Pro-drugs as
Novel Delivery Systems," Vol. 14 of the A.C.S. Symposium Series,
Edward B. Roche, ed., "Bioreversible Carriers in Drug Design,"
American Pharmaceutical Association and Pergamon Press, 1987, and
Judkins et al., Synthetic Communications 26(23):4351-4367, 1996,
each of which is incorporated herein by reference.
The term "sulfonyl," as used herein, represents --S(O).sub.2--.
By "thiol" is meant an --SH group.
Asymmetric or chiral centers may exist in the compounds of the
present invention. The present invention includes the various
stereoisomers and mixtures thereof. Individual stereoisomers of
compounds or the present invention may be prepared synthetically
from commercially available starting materials that contain
asymmetric or chiral centers or by preparation of mixtures of
enantiometic compounds followed by resolution well-known to those
of ordinary skill in the art. These methods of resolution are
exemplified by (1) attachment of a racemic mixture of enantiomers,
designated (+/-), to a chiral auxiliary, separation of the
resulting diastereomers by recrystallization or chromatography and
liberation of the optically pure product from the auxiliary, or (2)
direct separation of the mixture of optical enantiomers on chiral
chromatographic columns. Enantiomers are designated herein by the
symbols "R" or "S," depending on the configuration of substituents
around the chiral carbon atom, or are drawn by conventional means
with a bolded line defining a substituent above the plane of the
page in three-dimensional space and a hashed or dashed line
defining a substituent beneath the plane of the printed page in
three-dimensional space. If no stereochemical designation is made,
it is to be assumed that the structure definition includes both
stereochemical possibilities.
The invention provides several advantages. For example, as is noted
above, the invention provides an approach for treating inflammatory
bowel disease, which can be a very painful and debilitating
condition that is difficult to treat, and affects more than one
million people in the United States alone. The methods of the
invention can also be used to prevent or to treat other conditions
associated with TLR2 activation, as is discussed elsewhere herein.
Finally, the screening methods of the invention provide
straightforward approaches for identifying and characterizing
agents that can be used in the prevention and treatment of
TLR2-associated diseases and conditions.
Other features and advantages of the invention will be apparent
from the following detailed description and the claims.
DETAILED DESCRIPTION
The invention is based in part on our discovery that animals that
do not express Toll-like receptor 2 (TLR2) are protected from
dextran sulfate sodium (DSS) induction of colitis, a model for
inflammatory bowel disease (IBD). Based on this discovery, we
concluded that agents that block activation of TLR2 can be used to
treat or to prevent colitis and related diseases or conditions, as
well as other diseases or conditions characterized by activation of
TLR2. Accordingly, the invention provides compounds and methods for
preventing or treating diseases or conditions associated with
activation of TLR2, as well as methods for identifying agents that
decrease or inhibit activation of this receptor. The compounds and
methods of the invention are described in further detail, as
follows.
Preparation of TLR2 Inhibitors
A compound of formula I,
##STR00014## where X is --NHC(O)--; each of a and b is 1; W is
--C(O)N(R.sup.W1)(CH.sub.2).sub.cCH(OR.sup.W3)R.sup.W4, where c is
2, each of R.sup.W1 and R.sup.W4 is H, and each of U, V, and
R.sup.W5 is as defined above can be prepared by a sequence of
reactions shown in Scheme 1. Accordingly, a compound of formula III
is epoxidized to produce a compound of formula IV, and the epoxy
group reacted with a cyano group, which nucleophilically opens up
the epoxide to produce a compound of formula V. Methods of
preparing chiral epoxides from achiral starting materials are known
to those skilled in the art and such methods would produce a
compound of formula V of known configuration. Protection of the
hydroxyl group as the t-butyldiphenylsilyl ether, followed by
reduction of the cyano group with Raney nickel, produces a compound
of formula VII. Compounds of formula VIII can be prepared by
coupling a compound of formula VII with the L- or D-form of
N-Fmoc-serine. The hydroxyl group derived from the serine can be
subsequently reacted with phosphorylating agent IX, and the
intermediate phoshine oxidized with hydrogen peroxide to produce a
compound of formula X. The Fmoc protecting group can be selectively
removed with piperidine and the resulting amine acylated with an
acyl chloride or coupled to a compound containing a carboxyl group
in a reaction mediated by a carbodiimide or other suitable coupling
reagent. Subsequent removal of the silyl protecting group with
fluoride ion produces a compound of formula XI. Removal of the Boc
protecting group under acidic conditions (e.g., TFA or HCl/dioxane)
and acylation, reductive amination, or sulfonation of the resulting
amine results in a compound of formula XII. Treatment with a Pd(0)
catalyst removes the phosphonate allyl protecting group to produce
a compound of formula XIII.
##STR00015## ##STR00016##
Examples of compounds of Formula XIII are shown in Table 1, where
R.sup.1 is H, C2 has an (S)-configuration, and C3' is an
(R,S)-configurational mixture, unless otherwise specified.
TABLE-US-00001 TABLE 1 Compounds of formula XIII, where R.sup.1 is
H Compound No. U V R.sup.W4 ER810702 ##STR00017##
--(CH.sub.2).sub.18CH.sub.3 --(CH.sub.2).sub.6CH.sub.3 (R)-config.
ER811133 ##STR00018## --(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2).sub.12CH.sub.3 ER811134 ##STR00019##
--(CH.sub.2).sub.4CH.sub.3 --(CH.sub.2).sub.12CH.sub.3 ER811392
##STR00020## --(CH.sub.2).sub.2C.sub.6H.sub.5
--(CH.sub.2).sub.9CH.sub.3 ER811393 ##STR00021##
--(CH.sub.2).sub.18CH.sub.3 --(CH.sub.2).sub.12CH.sub.3 ER811394
##STR00022## --(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2).sub.9CH.sub.3
ER811395 ##STR00023## --(CH.sub.2).sub.18CH.sub.3
--(CH.sub.2).sub.9CH.sub.3 ER811254 ##STR00024##
--(CH.sub.2).sub.18CH.sub.3 --(CH.sub.2).sub.12CH.sub.3 ER811255
##STR00025## --(CH.sub.2).sub.18CH.sub.3
--(CH.sub.2).sub.12CH.sub.3 ER812011 ##STR00026##
--(CH.sub.2).sub.18CH.sub.3 --(CH.sub.2).sub.12CH.sub.3
Compounds of formula XV, where R.sup.W3 is a C.sub.1-20 alkyl,
C.sub.1-20 alkenyl, or C.sub.1-20 alkynyl group, can be prepared in
a manner analogous to the synthetic route shown in Scheme I via a
compound of formula XIV, which can be prepared by reacting a
compound of formula V with an alkyl halide or an alkyl mesylate
corresponding to R.sup.W3-Hal or R.sup.W3-OMs, respectively.
##STR00027##
Compounds of formula XV, where R.sup.W3 is a C.sub.1-21 acyl group,
can be prepared as shown in Scheme 2 via a compound of formula XI.
Compounds of formula XI can be N-deprotected under acidic
conditions and then reacted with a carboxylic acid using a coupling
reagent to give a protected phosphonate intermediate, which can be
subsequently treated with catalytic Pd(0), thereby removing the
allyl protecting group to produce the compound of formula XV.
##STR00028##
Examples of compounds of Formula XV, where R.sup.1 is H, R.sup.W3
is an acyl group, and C2 and C3' have the S- and R-configuration,
respectively, unless otherwise indicated are shown in Table 2.
TABLE-US-00002 TABLE 2 Compounds of formula XV, where R.sup.1 is H
Compound No. U V R.sup.W3 R.sup.W4 ER809834 ##STR00029##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.4CH.sub.3
--(CH.sub.2).- sub.6CH.sub.3 ER809835 ##STR00030##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER809836 ##STR00031##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.12CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER809841 ##STR00032##
--(CH.sub.2).sub.4CH.sub.3 --C(O)(CH.sub.2).sub.12CH.sub.3
--(CH.sub.2).- sub.6CH.sub.3 ER809842 ##STR00033##
--(CH.sub.2).sub.4CH.sub.3 --C(O)(CH.sub.2).sub.18CH.sub.3
--(CH.sub.2).- sub.6CH.sub.3 ER809845 ##STR00034##
--(CH.sub.2).sub.4CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2).- sub.6CH.sub.3 ER809846 ##STR00035##
--(CH.sub.2).sub.4CH.sub.3 --C(O)(CH.sub.2).sub.12CH.sub.3
--(CH.sub.2).- sub.6CH.sub.3 ER809950 ##STR00036##
--(CH.sub.2).sub.10CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER809951 ##STR00037##
--(CH.sub.2).sub.10CH.sub.3 --C(O)(CH.sub.2).sub.12CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER809959 ##STR00038##
--(CH.sub.2).sub.18CH.sub.3 --C(O)CH.sub.3
--(CH.sub.2).sub.6CH.sub.3 ER809960 ##STR00039##
--(CH.sub.2).sub.18CH.sub.3 --C(O)(CH.sub.2).sub.4CH.sub.3
--(CH.sub.2).- sub.6CH.sub.3 ER809964 ##STR00040##
--(CH.sub.2).sub.10CH.sub.3 --C(O)CH.sub.3
--(CH.sub.2).sub.6CH.sub.3 ER809965 ##STR00041##
--(CH.sub.2).sub.10CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER810676 ##STR00042##
--(CH.sub.2).sub.10CH.sub.3 --C(O)(CH.sub.2).sub.4CH.sub.3
--(CH.sub.2).- sub.12CH.sub.3 (R,S)-mixture at C3' ER810677
##STR00043## --(CH.sub.2).sub.10CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.12CH.sub.3
(R,S)-mixture at C3' ER810698 ##STR00044## --CH.sub.3
--C(O)(CH.sub.2).sub.18CH.sub.3 --(CH.sub.2).sub.6CH.sub.3 ER810699
##STR00045## --CH.sub.3 --C(O)(CH.sub.2).sub.12CH.sub.3
--(CH.sub.2).sub.6CH.sub.3 ER810701 ##STR00046## --CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2).sub.6CH.sub.3 ER808701
##STR00047## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 (R)-config. at C3'
--(CH.sub.2).sub.6CH.sub.3 ER808839 ##STR00048##
--(CH.sub.2).sub.12CH.sub.3 (R)-config. at C2
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2).sub.6CH.sub.3
Compounds of formula XV, where R.sup.W4 is H and R.sup.W3 is an
acyl group can be prepared as shown in Scheme 3. Accordingly,
N-Fmoc-L- or D-serine can be reacted with TBDPS-protected
propanolamine using an appropriate coupling agent, such as a
carbodiimide, to form a compound of formula XVI. The choice of L-
or D-serine determines the configuration at C2. The compound of
formula XVI can be subsequently reacted with Boc-phosphorylating
reagent IX, followed by oxidation with hydrogen peroxide to yield a
phosphonate of formula XVII. The silyl protecting group is removed
with tetrabutylammonium fluoride and the Boc protecting group
removed under acidic conditions to yield a compound of formula XIX.
Acylation, reductive amination, or sulfonation of the amine of a
compound of formula XIX gives a compound of formula XX. The
hydroxyl group of the compound of formula XX can be reacted with a
compound containing a carboxylic acid under coupling conditions to
give a compound of formula XXI. Removal of the Fmoc-protecting
group followed by coupling the resulting amine with a compound
containing a carboxylic acid under coupling conditions gives a
compound of formula XXIII. Removal of the allyl protecting group
using catalytic palladium tetrakistriphenylphosphine gives a
compound of formula XXIV.
##STR00049## ##STR00050##
Examples of compounds of Formula XXIV are shown in Table 3, where
R.sup.1 is H and C2 has an (S)-configuration.
TABLE-US-00003 TABLE 3 Compounds of formula XXIV, where R.sup.1 is
H Compound No. U V R.sup.W3 ER811203 ##STR00051## --CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 ER811211 ##STR00052##
--(CH.sub.2).sub.14CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
ER811212 ##STR00053## --(CH.sub.2).sub.10CH.sub.3
--C(O)(CH.sub.2).sub.14CH.sub.3 ER811213 ##STR00054##
--(CH.sub.2).sub.18CH.sub.3 --C(O)(CH.sub.2).sub.4CH.sub.3 ER811214
##STR00055## --(CH.sub.2).sub.4CH.sub.3
--C(O)(CH.sub.2).sub.18CH.sub.3 ER811219 ##STR00056##
--(CH.sub.2).sub.12CH.sub.3 --C(O)CH.sub.3 ER811220 ##STR00057##
--(CH.sub.2).sub.14CH.sub.3 --C(O)CH.sub.3 ER811221 ##STR00058##
--(CH.sub.2).sub.18CH.sub.3 --C(O)CH.sub.3 ER811228 ##STR00059##
--(CH.sub.2).sub.14CH.sub.3 --C(O)(CH.sub.2).sub.14CH.sub.3
ER811232 ##STR00060## --(CH.sub.2).sub.14CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 ER811233 ##STR00061##
--(CH.sub.2).sub.14CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
ER811237 ##STR00062## --(CH.sub.2).sub.14CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 ER811243 ##STR00063##
--(CH.sub.2).sub.14CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
ER811244 ##STR00064## --(CH.sub.2).sub.14CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 ER811249 ##STR00065##
--(CH.sub.2).sub.14CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
ER811250 ##STR00066## --(CH.sub.2).sub.14CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3
Compounds of formula I in which W is
--C(O)N(R.sup.W1)(CH.sub.2).sub.cCH(OR.sup.W3)(CH.sub.2).sub.eOR.sup.W5,
where each of c and e is 1, R.sup.W1 is H, R.sup.W5 is C.sub.1-21
acyl, and each of U and V is as defined above can be prepared by
starting with (R)- or (S)-2,2-dimethyl-1,3-dioxolane-4-methanol.
Conversion of the hydroxyl group to an amino group via mesylation,
displacement with azide, and reduction of the azide to an amine
gives a compound that, upon coupling to a serine derivative, gives
a compound of formula XXV. Removal of the acetonide protecting
group under acidic conditions and subsequent alkylation or
acylation of the resulting hydroxyl groups (exhaustively, resulting
in identical R.sup.W3 and R.sup.W5 groups, or selectively,
resulting in different R.sup.W3 and R.sup.W5 groups) gives a
compound of formula XXVI. This compound can be deprotected via a
hydrogenation reaction and carried forward using the synthetic
methodology previously described herein to give a compound of
formula XXVIII.
##STR00067##
Examples of a compound of formula XXVIII are ER811261 and
ER811245.
##STR00068##
For the preparation of compounds of formula I where W is H,
N-Fmoc-glycinol can be reacted with a compound of formula IX,
followed by subsequent reactions analogous to those shown in Scheme
1 to produce a compound of formula XXIX.
##STR00069##
Examples of compounds of formula XXIX, where R.sup.1 is H, are
shown in Table 4.
TABLE-US-00004 TABLE 4 Compounds of formula XXIX, where R.sup.1 is
H Compound No. U V ER811230 ##STR00070##
--(CH.sub.2).sub.14CH.sub.3 ER811231 ##STR00071##
--(CH.sub.2).sub.18CH.sub.3 ER811246 ##STR00072## ##STR00073##
ER811247 ##STR00074## ##STR00075## ER811248 ##STR00076##
##STR00077## ER811251 ##STR00078## ##STR00079## ER811252
##STR00080## ##STR00081## ER811253 ##STR00082## ##STR00083##
Compounds of formula I where W is --C(O)N(R.sup.W1)R.sup.W2 can be
prepared by coupling HN(R.sup.W1)R.sup.W2 to an appropriately
protected L- or D-serine analog to produce a compound of formula
XXX, followed by subsequent reactions analogous to those shown in
Scheme 1 to produce a compound of formula XXXI. Examples of
compounds of formula XXXI, where each of R.sup.1 and R.sup.W1 is H
and C2 has the S-configuration, unless indicated otherwise, are
shown in Table 4.
TABLE-US-00005 TABLE 4 Compounds of formula XXXI, where each of
R.sup.1 and R.sup.W1 is H ##STR00084## ##STR00085## Compound No. U
V R.sup.W2 ER811234 ##STR00086## --(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2).sub.9CH.sub.3 ER811132 ##STR00087##
--(CH.sub.2).sub.18CH.sub.3 --(CH.sub.2).sub.9CH.sub.3 ER811236
##STR00088## ##STR00089## --(CH.sub.2).sub.9CH.sub.3
Compounds of formula I where W is
--(CH.sub.2).sub.cO(CH.sub.2).sub.dCH(OR.sup.W3)R.sup.W4 can be
prepared as shown in Scheme 3 using the methodology described in
U.S. Patent Application Publication No. 20030153532 A1. A protected
serine analog is prepared by reaction L- or D-serine methyl ester
with ethyl benzimidate to produce the benzimidine of serine, which
is subsequently reduced with DIBAL to produce the compound of
formula XXXII. The hydroxyl group of XXXII can be O-alkylated with
the tosylate of formula XXXIII and the benzimidine removed by
treatment with refluxing 4M HCl to produce a compound of formula
XXXV. Amino group protection or N-acylation produces a compound of
formula XXXVI, which is subsequently reacted with a compound of
formula IX to produce a compound of formula XXXVII. By steps
analogous to those described above in Scheme 2, the compound of
formula XXXVII is transformed to a compound of formula XXXVIII.
Examples of compounds of formula XXXVIII are shown in Table 5,
where R.sup.1 is H and each of C2 and C3' has the R-configuration,
unless indicated otherwise.
##STR00090## ##STR00091##
TABLE-US-00006 TABLE 5 Compounds of formula XXXVIII, where
R.sup.1is H Compound No. U V R.sup.W3 R.sup.W4 ER811208
##STR00092## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811209 ##STR00093## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811210 ##STR00094## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER804469 ##STR00095## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER804529 ##STR00096## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER810625 ##STR00097## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811189 ##STR00098## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811197 ##STR00099## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811198 ##STR00100## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811199 ##STR00101## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811205 ##STR00102## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811215 ##STR00103## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811217 ##STR00104## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811222 ##STR00105## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811223 ##STR00106## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811225 ##STR00107## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811226 ##STR00108## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811258 ##STR00109## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER804283 ##STR00110## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER804335 ##STR00111## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER808263 ##STR00112## --(CH.sub.2).sub.12CH.sub.3 C2 has S-config.
--C(O)(CH.sub.2).sub.10CH.sub.3 C3 'has S-config.
--(CH.sub.2).sub.6CH.sub.3 ER808265 ##STR00113##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3 C3 'has
S-config. --(CH.sub.2).sub.6CH.sub.3 ER809050 ##STR00114##
--(CH.sub.2).sub.12CH.sub.3 C2 has S-config.
--C(O)(CH.sub.2).sub.10CH.sub.3 C3 'has S-config.
--(CH.sub.2).sub.6CH.sub.3 ER809388 ##STR00115##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER809406 ##STR00116##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808579 ##STR00117##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808580 ##STR00118##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808581 ##STR00119##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808584 ##STR00120##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808585 ##STR00121##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808586 ##STR00122##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808587 ##STR00123##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808588 ##STR00124##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808928 ##STR00125##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808929 ##STR00126##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808931 ##STR00127##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808932 ##STR00128##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808934 ##STR00129##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808936 ##STR00130##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808938 ##STR00131##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808939 ##STR00132##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808940 ##STR00133##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808941 ##STR00134##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808942 ##STR00135##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808943 ##STR00136##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808944 ##STR00137##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808945 ##STR00138##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808950 ##STR00139##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808951 ##STR00140##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808952 ##STR00141##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808953 ##STR00142##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808954 ##STR00143##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808955 ##STR00144##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808956 ##STR00145##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808957 ##STR00146##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808958 ##STR00147##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808959 ##STR00148##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808960 ##STR00149##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808961 ##STR00150##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808962 ##STR00151##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3 ER808963 ##STR00152##
--(CH.sub.2).sub.12CH.sub.3 --C(O)(CH.sub.2).sub.10CH.sub.3
--(CH.sub.2)- .sub.6CH.sub.3
ER808964 ##STR00153## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER808965 ##STR00154## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER808966 ##STR00155## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER808967 ##STR00156## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER808968 ##STR00157## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER808969 ##STR00158## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER808970 ##STR00159## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER808971 ##STR00160## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER808972 ##STR00161## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER808973 ##STR00162## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER808974 ##STR00163## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER808975 ##STR00164## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER808976 ##STR00165## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER808977 ##STR00166## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811238 ##STR00167## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811239 ##STR00168## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811240 ##STR00169## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811241 ##STR00170## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER811242 ##STR00171## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
ER118998 ##STR00172## --(CH.sub.2).sub.12CH.sub.3
--C(O)(CH.sub.2).sub.10CH.sub.3 --(CH.sub.2)- .sub.6CH.sub.3
Compounds of formula II, where a, b, R.sup.1, R.sup.2, R.sup.3,
R.sup.4, R.sup.5, U, X, and W are as previously defined herein, can
be prepared as shown in Scheme 4. Protected amino acids of formula
XXXIX can be coupled to appropriately protected amino acids of
formula XL to produce compounds of formula XLI. Removal of the
t-butyl ester under acid conditions gives a compound of formula
XLII, which can be subsequently coupled to a compound of formula
XLIII to produce a compound of formula II.
##STR00173##
Exemplary compounds of formula II include ER811204 and ER811195,
shown below.
##STR00174##
Compounds of formula I or II in which W is
--(CH.sub.2).sub.cO(CH.sub.2).sub.dCH(OR.sup.W3)R.sup.W4 can be
prepared by methods analogous to those described in U.S. Pat. No.
6,551,600. In one non-limiting example, a compound of formula XLIV,
where R.sup.W3 is C.sub.1-21 acyl and each of V and R.sup.W4 is,
independently, C.sub.1-20 alkyl, is coupled to
2-(4-allyloxybenzyl)malonic acid, mono allyl ester, which can be
prepared via the alkylation of the mono-allyl, mono-t-butyl ester
of malonic acid with the allyl ether of 4-bromomethylphenol,
followed by removal of the t-butyl protecting group under acidic
conditions. The resulting compound XLV is produced after removal of
the allyl protecting groups via treatment with a Pd(0)
catalyst.
##STR00175##
An example of a compound of formula XLV is ER808577.
##STR00176##
Compounds of formula I, where W is C(O)OR.sup.W2, can be prepared
by the coupling of protected serine analogs with alcohols using a
carbodiimide, or another suitable coupling reagent, as exemplified
in the reaction of N-Boc-O-benzyl-L-serine with a compound of
formula XLVI to produce a compound of formula XLVII.
##STR00177##
Selective removal of the Boc-protecting group under acidic
conditions, followed by N-acylation and subsequent removal of the
benzyl protecting group via hydrogenation gives a compound of
formula XLVIII. Phosphorylation, oxidation, and deprotection, as
previously described above, gives a compound of formula XLIX, which
can be functionalized at the free amine position by acylation,
reductive amination, or sulfonation to give, after removal of the
allyl protecting group, a compound of formula L.
##STR00178##
An example of a compound of formula L is ER808549
##STR00179##
Compounds of the invention can also incorporate peptide sequences.
Useful intermediates for incorporating peptides into compounds of
the invention are those that contain a carboxyl or amino group.
Intermediates that contain an amino group can be reacted with the
C-terminal carboxyl group of a protected peptide and intermediates
that contain a carboxyl group can be reacted with the amine
terminus of a protected peptide. A particularly useful intermediate
is that of formula LI, shown below, where X and W are as previously
defined herein.
##STR00180##
The peptide sequences can be synthesized by either solid or liquid
phase methods described and referenced in standard textbooks, or by
a combination of both methods. These methods are well known to
those skilled in the art, (see, for example, Bodanszky, In "The
Principles of Peptide Synthesis," Hafner, Rees, Trost, Lehn,
Schleyer, Zahradnik, Eds., Springer-Verlag, Berlin, 1984; Stewart
and Young, Solid Phase Peptide Synthesis, Pierce Chemical Co.,
Rockford, Ill., 1984).
All of the starting materials used in any of these methods are
commercially available from chemical vendors such as Aldrich,
Sigma, Nova Biochemicals, Bachem Biosciences, Advanced ChemTech,
and the like, or may be readily synthesized by known
procedures.
The reaction products are isolated and purified by conventional
methods, typically by solvent extraction into a compatible solvent.
The products may be further purified by column chromatography or
other appropriate methods, including medium pressure or high
pressure liquid chromatography.
During the synthesis of these compounds, the functional groups of
the amino acid derivatives used in these methods are protected by
blocking groups to prevent cross reaction during the coupling
procedure. Examples of suitable blocking groups and their use are
described in "The Peptides: Analysis, Synthesis, Biology," Academic
Press, Vol. 3 (Gross, E. & Meienhofer, J., Eds., 1981) and Vol.
9 (1987), the disclosures of which are incorporated herein by
reference.
A particularly useful support for the synthesis of peptide
sequences is 2-chlorotrityl resin. Peptide sequences prepared on
chlorotrityl resin can be further reacted in an on-resin reaction
with an intermediate used in the present invention that contains a
carboxyl group, such as, for example, a compound of formula XLI.
Alternatively, a peptide prepared on chlorotrityl resin can be
removed from the resin in the protected state, followed by reaction
of the carboxyl-terminus of the peptide with an intermediate of the
invention that contains an amine and subsequent removal of any
protecting groups used in the peptide synthesis. An example of a
compound of the present invention that contains a peptide sequence
is ER810625, shown below
##STR00181##
Compounds of formula LII:
##STR00182## where each of i, T, Z, R.sup.6, R.sup.7, R.sup.8, and
R.sup.9 is as defined above, can be prepared from an appropriately
protected amino acid which includes a sulfhydryl moiety as part of
the side chain, such as, for example, cysteine or homocysteine. The
sulfhydryl is reacted with an activated olefin to produce a
compound of formula LIII. After the Fmoc-protected amine LIII is
deprotected and manipulated in a manner analogous to that described
above for the elaboration of the X group of compounds of formula I
or II to give a compound of formula LIV, the carboxy terminus of a
compound of formula LIV can be functionalized as described herein
for the functionalization of other intermediates having a carboxyl
group to produce a compound of formula LII.
##STR00183##
An example of a compound of formula Ia is compound ER810675
##STR00184## Therapeutic Use of TLR2 Inhibitors
Agents that decrease or inhibit activation of TLR2 can be used to
prevent or to treat any of a number diseases or conditions that are
characterized by TLR2 activation. For example, the agents can be
used to prevent or to treat inflammatory bowel disease (IBD), such
as, for example, Crohn's disease or ulcerative colitis. Other
diseases or conditions that can be prevented or treated using the
methods of the invention include, for example, sepsis, periodontal
disease, mucositis, acne, cardiovascular disease, chronic
obstructive pulmonary disease, arthritis, cystic fibrosis,
bacterial-induced infections, viral-induced infections,
mycoplasma-associated diseases, post herpetic neuralgia,
ischemia/reperfusion injury, asthma, stroke, brain injury,
necrotizing enterocolitis, bed sores, leprosy, atopic dermatitis,
psoriasis, trauma, neurodegenerative disease, amphotericin
B-induced fever and nephritis, coronary artery bypass grafting, and
atherosclerosis.
Dosage levels of active ingredients in the pharmaceutical
compositions of the invention may be varied to obtain an amount of
the active compound(s) that achieves the desired therapeutic
response for a particular patient, composition, and mode of
administration. The selected dosage level depends upon the activity
of the particular compound, the route of administration, the
severity of the condition being treated, and the condition and
prior medical history of the patient being treated. For adults, the
doses are generally from about 0.01 to about 100 mg/kg, desirably
about 0.1 to about 1 mg/kg body weight per day by inhalation, from
about 0.01 to about 100 mg/kg, desirably 0.1 to 70 mg/kg, more
desirably 0.5 to 10 mg/kg body weight per day by oral
administration, and from about 0.01 to about 50 mg/kg, desirably
0.1 to 1 mg/kg body weight per day by intravenous administration.
Doses are determined for each particular case using standard
methods in accordance with factors unique to the patient, including
age, weight, general state of health, and other factors that can
influence the efficacy of the compound(s) of the invention.
It is not intended that the administration of a compound of the
invention to a mammal, including humans, be limited to a particular
mode of administration, dosage, or frequency of dosing. The present
invention contemplates all modes of administration, including oral,
intraperitoneal, intramuscular, intravenous, intraarticular,
intralesional, subcutaneous, or any other route sufficient to
provide a dose adequate to prevent or treat excess or undesired
TLR2 activity. One or more compounds or the invention may be
administered to a mammal in a single dose or multiple doses. When
multiple doses are administered, the doses may be separated from
one another by, for example, several hours, one day, one week, one
month, or one year. It is to be understood that, for any particular
subject, specific dosage regimes should be adjusted over time
according to the individual need and the professional judgment of
the person administering or supervising the administration of a
pharmaceutical composition that includes a compound of the
invention.
For clinical applications, a compound of the present invention may
generally be administered intravenously, subcutaneously,
intramuscularly, colonically, nasally, intraperitoneally, rectally,
buccally, or orally. Compositions containing at least one compound
of the invention that is suitable for use in human or veterinary
medicine may be presented in forms permitting administration by a
suitable route. These compositions may be prepared according to the
customary methods, using one or more pharmaceutically acceptable
adjuvants or excipients. The adjuvants comprise, inter alia,
diluents, sterile aqueous media, and various non-toxic organic
solvents. Acceptable carriers or diluents for therapeutic use are
well known in the pharmaceutical field, and are described, for
example, in Remington: The Science and Practice of Pharmacy (20th
ed.), ed. A. R. Gennaro, Lippincott Williams & Wilkins, 2000,
Philadelphia, and Encyclopedia of Pharmaceutical Technology, eds.
J. Swarbrick and J. C. Boylan, 1988-1999, Marcel Dekker, New York.
The compositions may be presented in the form of tablets, pills,
granules, powders, aqueous solutions or suspensions, injectable
solutions, elixirs, or syrups, and the compositions may optionally
contain one or more agents chosen from the group comprising
sweeteners, flavorings, colorings, and stabilizers in order to
obtain pharmaceutically acceptable preparations.
The choice of vehicle and the content of active substance in the
vehicle are generally determined in accordance with the solubility
and chemical properties of the product, the particular mode of
administration, and the provisions to be observed in pharmaceutical
practice. For example, excipients such as lactose, sodium citrate,
calcium carbonate, and dicalcium phosphate and disintegrating
agents such as starch, alginic acids, and certain complex silicates
combined with lubricants (e.g., magnesium stearate, sodium lauryl
sulfate, and talc) may be used for preparing tablets. To prepare a
capsule, it is advantageous to use lactose and high molecular
weight polyethylene glycols. When aqueous suspensions are used,
they may contain emulsifying agents that facilitate suspension.
Diluents such as sucrose, ethanol, polyethylene glycol, propylene
glycol, glycerol, chloroform, or mixtures thereof may also be
used.
For parenteral administration, emulsions, suspensions, or solutions
of the compositions of the invention in vegetable oil (e.g., sesame
oil, groundnut oil, or olive oil), aqueous-organic solutions (e.g.,
water and propylene glycol), injectable organic esters (e.g., ethyl
oleate), or sterile aqueous solutions of the pharmaceutically
acceptable salts are used. The solutions of the salts of the
compositions of the invention are especially useful for
administration by intramuscular or subcutaneous injection. Aqueous
solutions that include solutions of the salts in pure distilled
water may be used for intravenous administration with the proviso
that (i) their pH is adjusted suitably, (ii) they are appropriately
buffered and rendered isotonic with a sufficient quantity of
glucose or sodium chloride, and (iii) they are sterilized by
heating, irradiation, or microfiltration. Suitable compositions
containing a compound of the invention may be dissolved or
suspended in a suitable carrier for use in a nebulizer or a
suspension or solution aerosol, or may be absorbed or adsorbed onto
a suitable solid carrier for use in a dry powder inhaler. Solid
compositions for rectal administration include suppositories
formulated in accordance with known methods and containing at least
one compound of formula I or II.
Dosage formulations of a compound of the invention to be used for
therapeutic administration must be sterile. Sterility is readily
accomplished by filtration through sterile membranes (e.g., 0.2
micron membranes) or by other conventional methods. Formulations
typically are stored in lyophilized form or as an aqueous solution.
The pH of the compositions of this invention is typically between 3
and 11, more desirably between 5 and 9, and most desirably between
7 and 8, inclusive. While a desirable route of administration is by
injection such as intravenously (bolus and/or infusion), other
methods of administration may be used. For example, compositions
may be administered subcutaneously, intramuscularly, colonically,
rectally, nasally, or intraperitoneally in a variety of dosage
forms such as suppositories, implanted pellets or small cylinders,
aerosols, oral dosage formulations, and topical formulations such
as ointments, drops, and dermal patches. A compound of the
invention is desirably incorporated into shaped articles such as
implants, including but not limited to valves, stents, tubing, and
prostheses, which may employ inert materials such as synthetic
polymers or silicones, (e.g., Silastic, silicone rubber, or other
commercially available polymers). Such polymers can include
polyvinylpyrrolidone, pyran copolymer,
polyhydroxy-propyl-methacrylamide-phenol,
polyhydroxyethyl-aspartamide-phenol, or
polyethyleneoxide-polylysine substituted with palmitoyl residues.
Furthermore, a TLR2 inhibitor of the invention may be coupled to a
class of biodegradable polymers useful in achieving controlled
release of a drug, for example polylactic acid, polyglycolic acid,
copolymers of polylactic and polyglycolic acid, polyepsilon
caprolactone, polyhydroxy butyric acid, polyorthoesters,
polyacetals, polydihydropyrans, polycyanoacrylates, and cross
linked or amphipathic block copolymers of hydrogels.
A compound of the invention may also be administered in the form of
liposome delivery systems, such as small unilamellar vesicles,
large unilamellar vesicles, and multilamellar vesicles. Liposomes
can be formed from a variety of lipids, such as cholesterol,
stearylamine, or phosphatidylcholines. A compound of the invention
may also be delivered using antibodies, antibody fragments, growth
factors, hormones, or other targeting moieties to which the
compound molecules are coupled (e.g., see Remington: The Science
and Practice of Pharmacy, vide supra), including in vivo
conjugation to blood components of a compound of the formula I or
II, as described herein.
Identification of TLR2 Inhibitors
Pharmaceutical agents that can be used in the therapeutic methods
of the invention can be identified in screening methods. For
example, cell-based screening methods can be used, in which cells
expressing TLR2 are contacted with a candidate agent and the impact
of the agent on the activation of TLR2 in the cells is determined.
In one example of such a method, the effect of an agent on the
activation of TLR2 by a known ligand (e.g., a lipopeptide, such as
Pam3Cys-SKKKK (see below)) is determined. Agents that are found to
decrease or to block activation of the receptor by the ligand can
then be considered for further analysis and/or for use as TLR2
inhibitors in therapeutic methods. Activation of TLR2 in these
methods can be measured using, for example, a reporter system. For
example, cells used in the screening assay can include a reporter
gene (e.g., luciferase) that is under the control of a promoter
(e.g., the ELAM promoter) that is inducible by a signaling pathway
triggered by TLR2 activation (e.g., a signaling pathway that
induces expression of NF-KB). Additional details of an example of
such a method are provided below.
In addition to cell-based methods, candidate agents can be tested
in animal model systems. This may be desirable, for example, if an
agent has been found to have antagonist activity in a cell-based
assay or to bind to TLR2 in an in vitro assay (see below). For
example, in animal studies, test agents can be administered to an
animal model concurrently with a molecule known to activate TLR2
(e.g., lipopeptide), and the impact of the agent on a response in
the animal that is normally triggered by activation of the receptor
(e.g., cytokine induction) can be determined. Further, in vitro
methods can be used. For example, a candidate compound can be
assayed for whether it binds to TLR2 or a fragment of the receptor
that includes at least a portion of the ligand binding site. Such
assays can be carried out using, for example, columns or beads to
which the receptor or fragment is bound.
In addition to the methods described above, additional TLR2
antagonists can be identified in methods in which candidate
compounds are compared for TLR2 antagonist activity with any of the
TLR2 antagonists described herein. These methods can involve the
use of in vitro or in vivo methods, such as those described herein
(e.g., see above and Example 8). Further, in addition to being
compared for TLR2 antagonist activity, the candidate compounds can
be compared with TLR2 antagonists with respect to specificity for
TLR2 versus other receptors (e.g., TLR4), as described below.
Candidate compounds identified as having TLR2 antagonist activity
that is, for example, similar to or greater than the activity of
the antagonists described herein (and/or with similar or greater
levels of specificity for TLR2 versus TLR4) in these assays can be
tested further, for example, in appropriate animal model assays for
any of the diseases or conditions described herein, as well as in
human clinical studies. A candidate compound having TLR2 antagonist
activity is one that, for example, is found to have an IC.sub.50
for TLR2 in an assay such as that described in Example 8 of, e.g.,
less than 30 .mu.M (e.g., less than 25, 20, 15, 10, 5, 1, or 0.5
.mu.M).
Also included in the invention are compounds that are selective for
TLR2 over TLR4, as well as compounds that are dual antagonists
(i.e., antagonists of both TLR2 and TLR4). A compound that is
selective for TLR2 over TLR4 is one that has, for example, an IC50
value in a TLR2 antagonist assay, such as is described herein, that
is less than that found in a TLR4 antagonist assay, such as is
described herein. For example, the IC50 in the TLR2 assay can be at
least 5, 10, 25, or 50-fold less than the value for the same
compound tested in the TLR4 assay. Compounds that are dual
antagonists are those that have, for example, IC50 values that are
within a 5-fold range of one another using, e.g., the assays
described herein. Thus, dual antagonists include those that have
activities that are 1:5-5:1 with respect to one another (e.g., 1:4,
1:3, 1:2, 1:1, 2:1, 3:1, and 4:1). The invention also includes the
use of TLR2 antagonists such as those described herein in the study
of physiological and molecular pathways involved in or affected by
TLR2 activation (or inactivation).
Agents that can be screened using the methods of the invention
include, for example, compounds that are present in compound
libraries (e.g., random libraries), as well as analogs of known
TLR2 ligands (e.g., lipopeptides) that are modified to prevent
rather than activate TLR2. Further, peptides that correspond to the
binding site of TLR2 for its ligands, which can competitively
inhibit ligand binding to the binding site, can be tested. Further,
antibodies or antibody fragments to the ligand or the ligand
binding site of the receptor can be screened.
The following non-limiting examples are provided to further
describe various aspects and embodiments of the present
invention.
EXAMPLES
Example 1
Preparation of ER811133
Step 1. To a solution of 1-pentadecene (23.0 mL, 84.7 mmol) in DCM
(200 mL) at 0.degree. C. was added sodium bicarbonate (704 mg, 8.3
mmol) and mCPBA (58.48 g, 338.9 mmol) portion wise. The reaction
mixture was allowed to warm up to RT. After stirring at RT
overnight, saturated sodium sulfite solution (500 mL) was added to
the reaction mixture. The organic layer was dried over sodium
sulfate and concentrated in vacuo. Purification of the residue by
silica gel chromatography (10% EA/hexane) gave compound 1 (19.10 g,
96% yield).
##STR00185##
Step 2. To a solution of compound 1 (19.1 g, 84.4 mmol) in 90%
ethanol (200 mL) at RT was added potassium cyanide (15.38 g, 236.2
mmol). After stirring at RT for 20 hours, the reaction mixture was
filtered through sodium sulfate. Purification by silica gel
chromatography (30% EA/hexane) gave compound 2 (18.46 g, 86%
yield).
##STR00186##
Step 3. To a solution of compound 2 (18.46 g, 72.84 mmol) in DMF
(100.0 mL) at RT was added imidazole (9.9 g, 145.7 mmol) and
tert-butyldiphenylsilyl chloride (TBDPSCI) (28.4 mL, 109.3=mol).
The reaction was monitored by TLC (30% EA/hexane) until all the
starting material was consumed. Purification by silica gel
chromatography (20% EA/hexane) gave compound 3 (34.84 g, 97%
yield).
##STR00187##
Step 4. To a solution of Raney-Ni (3.0 mL slurry) in 2.0 M
NH.sub.3/MeOH (100 mL) was compound 3 (5.04 g, 10.2 mmol) and the
reaction mixture was hydrogenated at 50 psi for 20 hours. The
reaction mixture was filtered through Celite and washed with MeOH
to give compound 4 (4.81 g, 95%).
##STR00188##
Step 5. To a solution of compound 4 (4.81 g, 9.7 mmol) in DCM (50
mL) was added N-Fmoc-L-serine (4.76 g, 14.55 mmol) and EDC (3.72 g,
19.4 mmol) at -5.degree. C. The reaction mixture was allowed to
warm up to RT and stirred for 3 hours. Purification by silica gel
chromatography using 10-50% EA/hexane gave compound 5 (6.26 g, 80%)
as waxy solid.
##STR00189##
Step 6. To make phosphorylating reagent compound 6, to a solution
of distilled diisopropylamine (9.0 mL) in methylene chloride was
added tetrazole (4.51 g) at room temperature followed by stirring
for 1.5 hours. Allyl phosphorodiamidite (10) (20.5 mL) was added
dropwise at a 6.5 mL/hour rate followed by stirring for an
additional 3 hours. N-Boc-2-aminoethanol (10.36 g) in methylene
chloride (50 mL) was added to the above reaction mixture dropwise
at a 8.4 mL/hour rate followed by stirring for an additional 18
hours. The white suspension was filtered through Celite 545 with
two 20 mL washings with methylene chloride. The filtrate was
concentrated followed by the suspension and filtering of the
residue with hexanes (200 mL). The resulting hexanes filtrate was
concentrated to dry and azeotroped with 2,10-mL portions of toluene
to provide the crude product 6 (21.54 g) as an oil.
To a solution of compound 5 (6.26 g, 7.77 mmol) in DCM (70 mL) at
RT was added pyridinium trifluoroacetate (3.0 g, 15.55 mmol). The
above reaction solution was cooled to -20.degree. C. To the
reaction solution was added compound 6 (4.8 g, 14.0 mmol) using a
syringe. The reaction was kept at -10 to -20.degree. C. and
monitored by TLC (30% acetone/hexane) until compound 5 was
consumed. To the reaction mixture was added hydrogen peroxide (30%,
1.8 mL). The reaction mixture was allowed to warm up slowly and
stirred for 30 minutes at RT. To the solution was added sodium
thiosulfate (2.0 g) in water (20 mL). Compound 7 (8.7 g) was
isolated after aqueous workup and HPLC on a Biotage column (10-30%
acetone/hexane).
##STR00190##
Step 7. To a solution of compound 7 (2-83 g, 2.645 mmol) in DCM
(12.0 mL) was added piperidine (3.0 mL, 10.1 mmol) at -5.degree. C.
The mixture was stirred at -5.degree. C. for 3 0 minutes, followed
by warming to RT and monitoring by TLC. When the starting material
was consumed, the reaction mixture was concentrated to give crude
amine. To a solution of the crude amine in DCM (10 mL) was added
lauric acid (1.06 g, 5.29 mmol) and EDC (1.01 g, 5.29 mmol) at
0.degree. C. After stirring at 0.degree. C. for 110 minutes, the
reaction mixture was allowed to warm up to RT and stirred at RT
overnight. Aqueous workup, followed by concentration of the
organics and purification by chromatography gave compound 8 (2.45
g, 90% yield).
##STR00191##
Step 8. To a solution of compound 8 (1.5 g, 0.473 mmol) in THF (10
mL) at RT was added acetic acid (0.17 mL, 2.92 mmol) and
tetrabutylammonium fluoride (TBAF) (0.763 g, 2.92 mmol). After
stirring at RT for 70 hours, the reaction mixture was diluted with
water and extracted with EA. The combined organic solution was
washed with saturated sodium bicarbonate, dried and concentrated.
Purification by HPLC on a Biotage column with acetone/hexane gave
product compound 9 (0.93 7 g, 8 1% yield).
##STR00192##
Step 9. To compound 9 (138 mg, 0.175 mmol) was added 4.0 M
HCl/dioxane (2.0 mL) and the mixture stirred for 2 hours at RT. The
reaction mixture was concentrated and dried azeotropically with
toluene to give crude amine. To a solution of the crude amine in
DCM (5.0 mL) at 0.degree. C. was added
N-(3-indolacetyl)-L-isoleucine (55.4 mg, 0.192 mmol), HBTU (79.5
mg, 0.21 mmol) and DIPEA (90.3 mg, 0.70 mmol). The reaction was
allowed to warm to RT and stirred overnight. The reaction mixture
was loaded onto a 25 mm Biotage column and purified by HPLC,
eluting with 10, 20, 30, 40, and 50% acetone/hexane to give
compound 10 (72.0 mg, 43% yield).
##STR00193##
Step 10. To a solution of compound 10 (25.3 mg, 0.026 mmol) in THF
(1.0 mL) was added phenylsilane (29 .mu.L, 0.237 mmol),
triphenylphosphine (12.4 mg, 0.047 mmol) and
tetrakis(triphenylphosphine)palladium(0) (0.006 mg, 0.005 mmol) at
RT. After stirring at RT for 30 minutes, the crude mixture was
loaded onto 0.5 millimeter preparative TLC plate and eluted with
1:1 Magic (60:35:2:3 CHCl.sub.3/MeOH/AcOH/H.sub.2O)/DCM to give
ER811133 (18.3 mg, 75%).
##STR00194##
Example 2
Preparation of ER811212
Step 1. To a solution of Fmoc-Ser-OH (25.2 g, 77.0 mmol) in
dichloromethane (350 mL) at RT was added TBDPS protected
propanolamine hydrochloride (20.0 g, 51.75 mmol) followed by
triethylamine (14.5 mL, 104.1 mmol) and
1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide hydrochloride (EDC,
19.85 g, 103.5 mmol). Stirring overnight followed by aqueous
work-up and column chromatography gave compound 11 (10.38 g, 32%
yield).
##STR00195##
Step 2. To a solution of compound 11 (17.83 g, 28.63 mmol) in
dichloromethane (400 mL) at RT was added pyridinium
trifluoroacetate (12.26 g, 63.48 mmol). The reaction mixture was
cooled to -20.degree. C. and compound 6 (17.60 g, 50.51 mmol) was
added. After 2 hours, 30% hydrogen peroxide (14.5 mL, 140.3 mmol)
was added and the reaction was allowed to warm to RT. To the
reaction mixture was added a solution of sodium thiosulfate (17.2
g) in water (215 mL). Aqueous work-up and column chromatography
gave compound 12 (25.37 g, 100% yield).
##STR00196##
Step 3. To a solution of compound 12 (15.82 g, 17.85 mmol) in THF
(85 mL) at RT was added glacial acetic acid (2.1 mL, 36.68 mmol)
followed by TBAF (9.35 g, 35.8 mmol). After stirring for 8 hours,
aqueous work-up and column chromatography gave compound 13 (3.65 g,
31% yield).
##STR00197##
Step 4. To compound 13 (3.65 g, 5.64 mmol) at RT was added 4M HCl
in dioxane (21.5 mL, 86.1 mmol). After 20 minutes, solvent was
evaporated to give compound 14 (3.09 g, 93% yield).
##STR00198##
Step 5. To a solution of compound 14 (3.30 g, 5.64 mmol) in
dichloromethane (35 mL) at RT was added
N-(3-Indolylacetyl)-L-isoleucine (1.80 g, 6.24 mmol) followed by
DIEA (2.2 mL, 12.69 mmol) and HBTU (2.59 g, 6.83 mmol). After 2
hours, aqueous workup and chromatography gave compound 15 (3.74 g,
81% yield).
##STR00199##
Step 6. To a solution of compound 15 (204 mg, 0.25 mmol) in
dichloromethane (1.5 mL) at RT was added palmitic acid (125 mg,
0.49 mmol) followed by EDC (95 mg, 0.495 mmol) and DMAP (8.2 mg,
0.067 mmol). After stirring overnight at RT, aqueous work-up and
chromatography gave compound 16 (198 mg, 75% yield).
##STR00200##
Step 7. To compound 16 (65 mg, 0.061 mmol) at RT was added a
solution of 4.0 M piperidine in DMF (0.38 mL, 1.54 mmol). After 20
minutes, the solvent was evaporated to give compound 17 (50 mg,
100% yield).
##STR00201##
Step 8. To a solution of compound 17 (51.1 mg, 0.059 mmol) in
dichloromethane (0.48 mL) at RT was added lauric acid (17.2 mg,
0.086 mmol) followed by DIEA (24 .mu.L, 0.138 mmol) and HBTU (30.7
mg, 0.081 mmol). After stirring overnight, the solvent was
evaporated. Purification by column chromatography gave compound 18
(44.8 mg, 73% yield).
##STR00202##
Step 9. To a solution of compound 18 (45 mg, 0.044 mmol) in THF
(0.63 mL) at RT was added a 0.10 M solution of n-butylamine (0.450
mL, 0.045 mmol) followed by palladium tetrakistriphenylphosphine
(2.9 mg, 0.0025 mmol). After 2 hours, the solvent was evaporated.
Purification by column chromatography gave compound 19 (ER811212,
22.9 mg, 59% yield).
##STR00203##
Example 3
Preparation of ER811230
Step 1. To a solution of Fmoc-Glycinol (104.4 mg, 0.3685 mmol) in
dichloromethane (5 mL) at RT was added pyridinium trifluoroacetate
(155.1 mg, 0.8031 mmol). The reaction mixture was cooled to
-20.degree. C. and Boc-phosphorylating reagent compound 6 (212.1
mg, 1.652 mmol) was added. After 2 hours, 30% hydrogen peroxide
(0.170 mL, 1.665 mmol) was added and the reaction was allowed to
warm to RT. To the reaction mixture was added a solution of sodium
thiosulfate (240 mg in 3.2 mL water). Aqueous work-up and column
chromatography gave compound 20 (150 mg, 74% yield).
##STR00204##
Step 2. To compound 20 (150 mg, 0.27 mmol) at RT was added 4 M HCl
in dioxane (1.0 mL). After 20 minutes, solvent was evaporated to
give compound 21 (132 mg, 100% yield).
##STR00205##
Step 3. To compound 21 (132 mg, 0.27 mmol) in dichloromethane (1.5
mL) at RT was added N-(3-indolylacetyl)-L-isoleucine (85 mg, 0.29
mmol), followed by DIEA (0.1 mL, 0.57 mmol) and HBTU (117 mg, 0.31
mmol). After stirring for 2 hours at RT, aqueous workup and
chromatography gave compound 22 (141 mg, 71% yield).
##STR00206##
Step 4. To compound 22 (32 mg, 0.045 mmol) at RT was added a
solution of 4.0 M piperidine in DMF (0.260 mL). After 20 minutes,
the solvent was evaporated to give compound 23 (21 mg, 94%).
##STR00207##
Step 5. To a solution of compound 23 (21 mg, 0.04 mmol) in
dichloromethane (0.25 mL) at RT was added palmitic acid (13 mg,
0.05 mmol) followed by DIEA (16 .mu.L, 0.09 mmol) and HBTU (20 mg,
0.054 mmol). After stirring overnight, the solvent was evaporated.
Purification by column chromatography gave compound 24 (19 mg, 61%
yield).
##STR00208##
Step 6. To a solution of compound 24 (19 mg, 0.026 mmol) in THF
(0.4 mL) at RT was added a 0.1 M solution of n-butylamine (0.26 mL,
0.026 mmol) followed by palladium tetrakistriphenylphosphine (1.7
mg, 0.0015 mmol). After 2 hours, the solvent was evaporated.
Purification by column chromatography gave compound ER811230 (13.0
mg, 73% yield).
##STR00209##
Example 4
Preparation of ER8 11261
Step 1. To a solution of
(S)-(+)-2,2-dimethyl-1,3-dioxolane-4-methanol (1.36 g, 10.3 mmol)
in dichloromethane (5 mL) was added triethylamine (4.30 mL, 30.9
mmol). The mixture was cooled with an ice/water bath and
methanesulfonyl chloride (2.40 mL, 30.9 mmol) was added. The
reaction was quenched by addition of saturated NaHCO.sub.3 solution
(aqueous) after stirring overnight. The aqueous phase was extracted
with one portion of dichloromethane; combined organic phase was
washed with water, dried with anhydrous sodium sulphate and
concentrated. Crude compound 25 was used without purification for
next step.
##STR00210##
Step 2. To a solution compound 25 from step 1 in DMF (10 mL) was
added a solution of NaN.sub.3 (2.0 g, 31 mmol) in water (5 mL) and
the reaction mixture was heated to 80.degree. C. After stirring 18
hours, reaction mixture was allowed to cool to RT and a brine
solution was added. The mixture was extracted with three portions
of diethyl ether and organic phase was concentrated to 1/2 volume
by rotary evaporation, washed with two portions of water, followed
by brine, dried with anhydrous sodium sulphate and concentrated.
Crude compound 26 (1.84 g) was used without purification for the
next step.
##STR00211##
Step 4. A solution of crude compound 26 (1.84 g),
triphenylphosphine (3.51 g, 13.4 mmol) in THF (20 mL) and water
(0.80 mL) was stirred at RT overnight. The solvent was evaporated
under reduced pressure and hexane was added to the residue. The
resulting solid was removed by filtration and this process of
adding hexane and filtration was repeated several times. The crude
produce was passed through a short column of silica gel (100% EA,
30% isopropyl alcohol/EA) to afford compound 27 (279 mg).
##STR00212##
Step 5. A mixture containing compound 27 (268 mg, 2.04 mmol), EDC
(589 mg, 3.07 mmol), DMAP (50 mg, 0.407 mmol), and
O-benzyl-N-acetyl-D-serine (725 mg, 3.07 mmol) in dichloromethane
(2 mL) was stirred at RT. After stirring overnight, the reaction
mixture was concentrated and purification by flash chromatography
gave compound 28 (359 mg).
##STR00213##
Step 6. A solution of acetic acid in water (3.50 mL, 5:1) was added
to compound 28 (343 mg, 0.979 mmol). After stirring at RT
overnight, the reaction was heated to 40.degree. C. After 1.5
hours, the reaction mixture was concentrated and azeotroped with
toluene. Purification by flash chromatography afforded compound 29
(289 mg, 95%).
##STR00214##
Step 7. A mixture of compound 29 (265 mg, 0.854 mmol), palmitic
acid (679 mg, 2.65 mmol), EDC (507 mg, 2.65 mmol), and DMAP (32.5
mg, 0.265 mmol) was stirred at RT for 18 hours. Aqueous work-up and
chromatography gave compound 30 (591 mg).
##STR00215##
Step 8. To a flask containing compound 30 in ethyl acetate (30 mL)
was added Pearlman's catalyst and the reaction mixture was stirred
under a hydrogen atmosphere (using hydrogen filled balloon). After
2 hours the reaction mixture was filtered through Celite and
concentrated to give compound 31 (344 mg). The crude product was
used in the next reaction without purification.
##STR00216##
Step 9. To a solution of compound 31 (51.7 mg, 0.0742 mmol) in DCM
(0.7 mL) at RT was added pyridinium trifluoroacetate (31.5 mg,
0.163 mmol). The above reaction solution was cooled to -20.degree.
C. To the reaction solution was added phosphorylating reagent
compound 6 (46.0 mg, 0.131 mmol). The reaction was kept at -10 to
-20.degree. C. and monitored by TLC (30% acetone/hexane) until
compound 31 was consumed. To the reaction mixture was added
hydrogen peroxide (30%, 18 .mu.L). The reaction mixture was allowed
to warm up slowly and stirred for 30 minutes at RT. After the
addition of sodium thiosulfate (20 mg), compound 32 was isolated
after aqueous workup and chromatography.
##STR00217##
Step 10. To compound 32 (42.3 mg, 0.044 mmol) was added 4.0 M
HCl/dioxane (132 .mu.L) and the mixture stirred for 2 hours at RT.
The reaction mixture was concentrated and dried azeotropically with
toluene to give crude amine. To a solution of the crude amine in
DCM (200 .mu.L) at 0.degree. C. was added
N-(3-(5-benzyloxy)indolacetyl)-L-isoleucine (20.0 mg, 0.0485 mmol),
HBTU (22.0 mg, 0.0582 mmol) and DIPEA (31.0 .mu.L, 0.176 mmol). The
reaction was allowed to warm to RT and stirred overnight. Aqueous
workup and chromatography gave compound 33 (72.0 mg, 43%
yield).
##STR00218##
Step 11. To a solution of compound 33 (25.3 mg, 0.026 mmol) (10.2
mg, 0.00825 mmol) in THF (300 .mu.L) was added phenylsilane (10
.mu.L, 0.08 mmol), triphenylphosphine (4.1 mg, 0.015 mmol) and
tetrakis(triphenylphosphine)palladium(0) (0.002 mg, 0.0015 mmol) at
RT. After stirring at RT for 30 minutes, the crude mixture was
loaded onto 0.5 millimeter preparative TLC plate and eluted with
1:1 Magic (60:35:2:3 CHCl.sub.3/MeOH/AcOH/H.sub.2O)/DCM to give
ER811261 (5.5 mg).
##STR00219##
Example 5
Preparation of ER808977
To a solution of compound 34 (100 mg, 0.11 mmol, prepared as
described in U.S. Pat. No. 6,290,973) in DCM (2.0 mL) was added
triethylsilane (87 .mu.L, 0.545 mmol) and trifluoroacetic acid (84
.mu.L, 1.1 mmol) at 0.degree. C. The resulting solution was at RT
until starting material was consumed (ca. 2 h). The reaction
mixture was then concentrated to give the TFA salt. The residue was
redissolved in DCM and washed with aqueous saturated sodium
bicarbonate solution (.times.2), and dried over Na.sub.2SO.sub.4.
Evaporation gave the crude amine free base, which was used
immediately without further purification. To a solution of
N-(3-indolacetyl)-L-isoleucine (R.sup.UCO.sub.2H, 29 mg, 0.1 mmol)
in DMF was added triethylamine and polymer supported carbodiimide.
The reaction mixture was shaken gently at RT for 2 hours then
treated with a solution of the aforementioned crude amine free base
in DMF. The reaction mixture was shaken gently at RT overnight.
Filtration, followed by concentration in vacuo, gave the crude
coupled product. The allyl protecting group(s) was removed by
treatment with a catalytic amount of palladium
tetrakistriphenylphosphine in THF in the presence of n-butylamine
for 2 hours. Purification by flash chromatography gave compound
ER808977. Other suitable protected amino acids can be used in place
of N-(3-indolacetyl)-L-isoleucine to react with (11) or an analog
thereof to produce the compounds shown in Table 5.
Aromatic intermediates that are useful for the preparation of the
compounds of the invention, such as, for example, compounds that
contain a substituted indole moiety, which are not commericially
available can be prepared from aromatic compounds that contain a
leaving group such as, for example, a halogen or a triflate. These
compounds can be reacted with a palladium catalyst/ligand system
(such as, for example, Pd(PPh.sub.3).sub.4, Pd(PtBu.sub.3).sub.4,
Pd[P(Me)(tBu.sub.2)].sub.4, PdCl.sub.2(PPh.sub.3).sub.2,
PdCl.sub.2(dppf).sub.2, Pd.sub.2(dba).sub.3BINAP, or
Pd.sub.2(dba).sub.3P(o-tol).sub.3) in the presence of a base and an
organometallic compound, such as for example, a compound with a
--B(OH).sub.2 or --B(OAlkyl).sub.2 group (Suzuki reaction),
--Mg-Hal group (Kumada reaction), --Zn-Hal group (Negishi
reaction), --Sn(Alkyl).sub.3 group (Stille reaction),
--Si(Alkyl).sub.3 group (Hiyama reaction), --Cu-Hal group,
--ZrCp.sub.2Cl group, or --AlMe.sub.2 group (see Fu and Littke,
Angew. Chem. Int. Ed. 41:4176-4211, 2002 for a review of
palladium-catalyzed cross-coupling reactions).
##STR00220##
Example 6
Preparation of 804469
Step 1. To a solution of mono-tert-butyl malonate (1.6 g, 10 mmol)
in DMF (20 mL) was added cesium carbonate (3.42 g, 10.5 mmol)
followed by allyl bromide (1.33 g, 11 1 mmol). The reaction was
stirred at RT for 16 hours, diluted with EA and worked up with
water and brine. The organic solution was dried over NaSO.sub.4 and
concentrated. The residue was purified by chromatography (silica
gel eluted with EA-hexanes) to give compound malonic acid,
monoallyl, mono-t-butyl ester (1.873 g, 93%).
##STR00221##
Step 2. To a solution of compound malonic acid, monoallyl,
mono-t-butyl ester (1.873 g, 9.27 mmol) in DMF (10 mL) was added
sodium hydride (400 mg, 60%, 10 mmol). The reaction was stirred at
RT for 20 minutes and followed by addition of
6-tert-butyldiphenylsiloxyl-1-bromohexane (3.69 g, 8.79 mmol). The
reaction was stirred at RT for 16 hours, followed by dilution with
EA and aqueous workup. The organics were dried over sodium sulfate
and concentrated. The residue was purified by chromatography
(silica gel eluted with EA-hexanes) to give compound 35 (3.947 g,
83% yield).
##STR00222##
Step 3. To compound 35 (2.10 g, 3.90 mmol) was added a solution of
tetrabutylammonium fluoride in THF (1 M, 5 mL). The reaction was
stirred at RT for 2 hours. The solvents were evaporated in vacuo
and the residue purified by chromatography (silica gel eluted with
EA-hexanes) to give 2-(6-hydroxylhexyl)malonic acid, monoallyl,
mono-t-butyl ester (460 mg, 39% yield).
##STR00223##
Step 4. To a solution of 2-(6-hydroxylhexyl)malonic acid,
monoallyl, mono-t-butyl ester (220 mg, 0.733 mmol) in
dichloromethane (3.5 mL) was added tetrazole (154 mg, 2.20 mmol).
The reaction was stirred at RT for 20 minutes followed by addition
of diallyl diisopropylphosphoramidite (270 mg, 1.10 mmol). The
reaction was stirred at room for 1 hour. The reaction mixture was
cooled to 0.degree. C. and 4 mL of THF was added followed by a
solution of Oxone (901 mg, 1.466 mmol) in water (4 mL). Stirring
was continued for 30 minutes, during which time the reaction was
allowed to warm up to RT. The reaction mixture was quenched by
adding aqueous sodium thiosulfate and sodium bicarbonate. The two
phases were separated and the aqueous phase was extracted with
dichloromethane. The organic phases were combined, washed with
brine, dried over sodium sulfate, and concentrated under vacuum.
The residue was purified by chromatography (silica gel eluted with
EA-hexanes) to give compound 36 (169 mg, 50% yield).
##STR00224##
Step 5. To a solution of compound 36 (77 mg, 0.167 mmol) in
dichloromethane (0.2 mL) was added triethylsilane (97 mg, 0.836
mmol) followed by trifluoroacetic acid (1 mL). The reaction was
stirred at RT for 1 hour and the volatiles were evaporated. The
residue was azeotroped with toluene twice and the crude product,
compound 37 (77 mg, 0.167 mmol), was used directly in the next
step.
##STR00225##
Step 6. To a solution of compound 38 (90 mg, 0.112 mmol, see U.S.
Pat. No. 6,290,973) and compound 37 (77 mg crude, 0.167 mmol) in
DMF (2.4 mL) was added
1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (38 mg,
0.2 mmol), 1-hydroxybenzotriazole hydrate (23 mg, 0.167 mmol), and
triethylamine (20 mg, 0.2 mmol). The reaction was stirred at RT for
16 hours, diluted by EA, washed with water, brine, and dried over
sodium sulfate. Purification by chromatography silica gel eluted
with MeOH-DCM) gave compound 39 (133 mg, 95%).
##STR00226##
Step 8. To a solution of compound 39 (133 mg, 0.112 mmol) and
phenylsilane (72 mg, 0.666 mmol) in CHCl.sub.3 (11 mL) cooled at
0.degree. C. in an ice-water bath was added
tetrakis(triphenylphosphine)-palladium(0) (192 mg, 0.166 mmol). The
reaction mixture was stirred at 0.degree. C. for 1.5 hours and
quenched by adding CHCl.sub.3:MeOH:H.sub.2O 2:3:1 (1 mL). The
reaction mixture was purified by a DEAE column eluted with 0.05-0.1
M NH.sub.4OAc in CHCl.sub.3:MeOH:H.sub.2O 2:3:1. Further
purification by HPLC (0/100 to 100/0A/B in 60 minutes.
A:Hexane-isopropanol-H.sub.2O 370-540-90, B: H.sub.2O-isopropanol
1-10) gave (ER804469) (35 mg, 30% yield).
##STR00227##
Example 7
Preparation of 811195
Step 1. To a solution of glycine t-butyl ester (511 mg, 3.05 mmol)
and N-(3-indolylacetyl)-L-isoleucine (878 mg, 3.05 mmol) in DMF/DCM
(5 mL/5 mL) at RT was added DIPEA (2.1 mL, 12.2 mmol) and HBTU
(2.31 g, 6.10 mmol). After stirring for 1 hour, the reaction
mixture was diluted with EA and washed with water (2.times.) and
brine (1.times.). Purification by silica gel chromatography gave
compound 40 (830 mg, 68% yield).
##STR00228##
Step 2. To compound 40 (83.0 mg, 0.207 mmol) was added 4 M HCl
solution in dioxane (2 mL, 8 mmol). After stirring at RT for 18
hours, the reaction mixture was concentrated and azeotroped with
THF/Toluene to give compound 41 (93.3 mg, overweight).
##STR00229##
Step 3. To a solution of compound 42 (2.05 g, 3.20 mmol, prepared
as described in U.S. Pat. No. 6,290,973) in DCM (20 mL) at
0.degree. C. was added DMAP (78.0 mg, 0.64 mmol) and Et.sub.3N (892
.mu.L, 6.40 mmol), followed by MsCl (496 .mu.L, 6.40 mmol). The ice
bath was removed and the reaction mixture stirred at RT for 30
minutes. The reaction mixture was cooled to 0.degree. C. and
quenched with 0.5 N HCl. The aqueous layer was extracted with DCM
(2.times.). Purification by silica gel chromatography gave compound
43 (2.30 g, 100% yield).
##STR00230##
Step 4. To a solution of compound 43 (2.30 g, 3.20 mmol) in DMF (10
mL) at RT was added NaN.sub.3 (624 mg, 9.6 mmol). The reaction
mixture was heated at 1 00.degree. C. for a total of 20 hours.
After cooling to RT, the reaction mixture was diluted with EA and
washed with water and brine. Purification by silica gel
chromatography gave compound 44 (1.24 g, 58% yield).
##STR00231##
Step 5. To a solution of compound 44 (1.24 g, 1.86 mmol) in MeOH
(20 mL) at RT was added Pd/C (200 mg). The reaction mixture was
stirred under an atmosphere of hydrogen for 4 hours. After
filtration through Celite, the crude product was purified by silica
gel chromatography to give compound 45 (793 mg, 64% yield).
##STR00232##
Step 6. To a solution of compound 45 (102 mg, 0.16 mmol) and
Boc-L-serine (39 mg, 0.19 mmol) in DCM (1 mL) was added EDC (46 mg,
0.24 mmol). After stirring for 20 hours at RT, the reaction mixture
was concentrated and purified on prep TLC to give compound 46 (113
mg, 86% yield).
##STR00233##
Step 7. To compound 46 (113 mg, 0.137 mmol) was added 4.0 N HCl
(1.5 mL, 6.0 mmol). After stirring at RT for 2.5 h, the reaction
mixture was concentrated and azeotroped with toluene (2.times.) to
give compound 47 (108 mg, 100% yield).
##STR00234##
Step 8. To a solution of compound 47 (108 mg, 0.137 mmol) and
compound 41 (79.0 mg, 0.207 mmol) in DMF (1.0 mL) was added EDC (28
mg, 0.411 mmol), HOBT (48 .mu.L, 0.207 mmol), and DIPEA (48 mL,
0.274 mmol). After stirring at RT for 17 hours, the reaction
mixture was diluted with EA and washed with water and brine.
Purification by silica gel chromatography gave compound 48 (13.7
mg, 9.5% yield).
##STR00235##
Step 9. To the solution of compound 48 (13.7 mg, 0.013 mmol) in
pyridine (1.0 mL) was added Py-SO.sub.3 (excess). After stirring at
RT for 24 hours, the reaction mixture was filtered through cotton,
concentrated and azeotroped with toluene. Purification by silica
gel chromatography gave ER811195 (8.6 mg, 58% yield).
##STR00236##
Example 7
Preparation of 809266
Step 1. To a solution of 3(R)-1,3-dihydroxydecane (6.39 g, 36.6
mmol) in DCM (40 mL) at 0.degree. C. was added DMAP (0.447 g, 3.66
mmol), triethylamine (10.2 mL, 73.2 mmol) and TBDPSCI (10.7 mL,
40.4 mmol). After stirring at RT for 20 hours, the reaction mixture
was diluted with DCM, washed with 0.5 N HCl and saturated
bicarbonate. The combined organic solution was dried with sodium
sulfate and concentrated to give crude compound 49 (about 16.5 g),
which was used as is.
##STR00237##
Step 2. To a solution of crude compound 49 (5.35 g, about 13.0
mmol) in anhydrous THF (45 mL) at 0.degree. C. was added
triphenylphosphine (10.3 g, 38.9 mmol), 4-nitrobenzonic acid (6.5
g, 38.9 mmol) and DEAD (6.1 mL, 38.9 mmol). After stirring at
0.degree. C. for 60 minutes, the reaction mixture was kept at
-20.degree. C. in a freezer overnight. Purification on a Biotage
column using EA/Hexane gave compound 50 (5.77 g, 79% yield).
##STR00238##
Step 3. To a solution of compound 50 (5.77 g, 10.3 mmol) in 1:1
THF:MeOH (120 mL) at RT was added potassium carbonate (1.72 g,
12.33 mmol) in one portion. Stirring was continued at RT for 4
hours. Purification on a Biotage column using EA/Hexane gave
compound 51 (3.26 g, 77% yield).
##STR00239##
Step 4. To a solution of compound 51 (3.26 g, 7.9 mmol) in
anhydrous DCM (20 mL) at 0.degree. C. was added DMAP (0.193 g, 1.5
mmol), and triethylamine (2.2 mL, 15.2 mmol), followed by the slow
addition of methanesulfonyl chloride (1.22 mL, 15.2 mmol). After
stirring at 0.degree. C. for 30 minutes, the reaction mixture was
allowed to warm up to RT and stirred for an additional 30 minutes.
The reaction mixture was diluted with 0.5 M HCl (50 mL) and
extracted with DCM (3.times.40 mL). The organic layer was washed
with saturated sodium bicarbonate solution, dried with MgSO4,
filtered, and concentrated to give compound 52 (4.0 g), which was
used directly as is in the next reaction.
##STR00240##
Step 5. To a solution of compound 52 (3.6 g, 7.34 mmol) in DMF (30
mL) was added sodium azide (1.45 g, 22.3 mmol) at RT. The reaction
mixture was heated in an oil bath at 140.degree. C. overnight.
After dilution with water and extraction with EA, the combined
organic layers were dried with MgSO.sub.4 and concentrated.
Purification on a Biotage column using 10% EA/Hexane gave compound
53 (2.27 g, 70% yield).
##STR00241##
Step 6. To a solution of compound 53 (2.25 g, 5.15 mol) in
anhydrous THF (20 mL) at RT was added tetrabutylammonium fluoride
(4.04 g, 15.44 mmol) in one portion. The mixture was stirred at RT
for 90 minutes and diluted with water/EA. The aqueous layer was
extracted with EA (3.times.20 mL), washed with 0.1 N HCl (20 mL)
and saturated sodium bicarbonate (20 mL). Flash chromatography with
Biotage 40 mm column (30% EA/Hexane) gave compound 54 (1.02 g, 99%
yield).
##STR00242##
Step 7. To a solution of compound 54 (1.01 g, 5.1 mmol) in DCM
(20.0 mL) at 0.degree. C. was added DMAP (45.0 mg, 0.37 mmol),
triethylamine (1.4 mL, 10.1 mmol), and methanesulfonyl chloride
(0.78 mL, 10.1 mmol). The resulting mixture was stirred for 2 hours
at 0.degree. C. and overnight at RT. The reaction mixture was
diluted with water, extracted with EA (3.times.20 mL), dried with
MgSO.sub.4, filtered, and concentrated. Flash chromatography using
Biotage 25 mm column (20% EA/Hexane) gave compound 55 (1.27 g, 90%
yield).
##STR00243##
Step 8. To a suspension of potassium tert-butoxide (0.98 g, 8.73
mmol) in THF (5.0 mL) at 0.degree. C. was added a solution of
benzimidate compound 56 (from L-serine methyl ester) (1.5466 g,
8.73 mmol) in THF (10.0 mL) using a syringe pump at a rate of 0.24
mL/minute. The resulting reaction mixture was stirred for 1 hour at
0.degree. C. To the mixture was added a solution of compound 55
(1.21 g, 4.36 mmol) in THF (10.0 mL) via syringe pump at a rate of
0.24 mL/minute. The reaction mixture was stirred for 48 hours.
Flash chromatography (30% EA/Hexane) gave compound 57 (0.73 g, 46%
yield).
##STR00244##
Step 9. To a solution of compound 57 (0.93 g, 2.6 mmol) in MeOH (10
mL) at RT was added 2 N HCl (7.8 mL, 15.6 mmol). The reaction
mixture was heated for 2 hours at 90.degree. C. After the reaction
mixture was cooled down to RT, a solution of 7.4 N NaOH (4.2 mL,
31.2 mmol) was added to the reaction mixture at RT. The reaction
mixture was heated for 4 hours at 90.degree. C., cooled to RT, left
standing overnight. The reaction mixture was extracted with DCM
(3.times.50 mL) and the combined organics concentrated to give
crude compound 58 (0.81 g), which was use for next step without
further purification.
##STR00245##
Step 10. To a solution of compound 58 (0.72 g) in THF (10 mL) at RT
was added saturated sodium bicarbonate (10 mL). To the resulting
mixture was added myristoyl chloride (0.63 mL, 2.24 mmol) at
0.degree. C. Stirring was continued for 20 minutes at 0.degree. C.
The reaction mixture was diluted with water, extracted with DCM
(3.times.10 mL), dried and concentrated. Purification by flash
chromatography (30% EA/Hexane) gave compound 59 (0.92 g, 76% yield
for 2 steps).
##STR00246##
Step 11. To a solution of compound 59 (0.92 g) in dichloromethane
(25 mL) at RT was added pyridinium trifluoroacetate (803 mg). The
reaction mixture was cooled to -20.degree. C. and
Boc-phosphorylating reagent compound 6 (1.10 g) was added. After 2
hours, 30% hydrogen peroxide (0.88 mL) was added and the reaction
was allowed to warm to RT. To the reaction mixture was added a
solution of sodium thiosulfate (1.24 g in 20 mL water). Aqueous
work-up and column chromatography gave compound 60 (0.619 g, 47%
yield).
##STR00247##
Step 12. To a solution of compound 60 (0.29 g, 0.38 mmol) in 20/1
THF:H.sub.2O (5.5 mL) at RT was added triphenylphosphine (0.18 g,
0.69 mmol). The resulting mixture was stirred at RT overnight,
concentrated, and dried azeotropically with toluene to give
compound 61, which was used for next step without purification.
##STR00248##
Step 13. To a solution of crude compound 61 (0.50 g) in anhydrous
DCM (5.0 mL) at 0.degree. C. was added lauric acid (0.14 g, 0.70
mmol), EDC (0.13 g, 0.70 mmol), and HOBt (0.094 g, 0.70 mmol). The
resulting mixture was stirred at RT until compound 61 was consumed.
Flash chromatography gave compound 62 (0.122 g, 39% yield for 2
steps).
##STR00249##
Step 14. To a solution of compound 62 (0.122 g, 0.135 mmol) in DCM
was added triethylsilane (108 .mu.L, 0.67 mmol) and TFA (104 .mu.L,
1.35 mmol) at 0.degree. C. The resulting mixture was stirred at RT
and monitored by TLC (10% MeOH/DCM) until compound 62 was consumed.
The reaction mixture was diluted with water and saturated sodium
bicarbonate, extracted with DCM (3.times.20 mL), dried with sodium
sulfate, and concentrated to give the crude amine compound 63,
which was used directly in the next reaction without further
purification.
##STR00250##
Step 15. To a solution of compound 63 (55.4 mg, 0.069 mmol) and
2-(4-allyloxybenzyl)malonic acid, mono allyl ester (60.1 mg, 0.207
mmol) in DCM (3.0 mL) was added EDC (26.3 mg, 0.138 mmol), HOBt
(18.6 g, 0.138 mmol), and triethylamine (72.2 .mu.L, 0.41 mmol) at
-5.degree. C. The mixture was stirred at RT overnight, diluted with
saturated bicarbonate, extracted with DCM (3.times.20 mL), dried
with sodium sulfate, and concentrated. Flash chromatography on a 25
mm Biotage column (40% Acetone/Hexane) gave compound 64 (52 mg, 69%
yield).
##STR00251##
Step 16. To a solution of compound 64 (51.7 mg, 0.048 mmol) in THF
was added phenylsilane (53.4 .mu.L, 0.433 mmol), triphenylphosphine
(22.7 mg, 0.086 mmol), and tetrakis(triphenylphosphine)palladium(0)
(16.7 mg, 0.014 mmol) at RT. The reaction mixture was stirred at RT
for 60 minutes and loaded onto DEAE, eluted with
2:3:1/CHCl.sub.3:MeOH:H.sub.2O to remove the palladium catalyst,
followed by column elution with 0.01 M, 0.02 M NH.sub.4OAC in
2:3:1. Product fractions were collected and diluted with an equal
volume of DCM. The organic solution was concentrated to give
ER809266 (32.8 mg, 71% yield).
##STR00252##
ER809265 and ER809267 can be prepared by a similar sequence of
reactions, with the exception that the configuration of the chiral
center of compound 49 is not inverted before mesylation and azide
displacement.
##STR00253##
Example 8
Identification of TLR2 and TLR4 Antagonists
Materials
Either Pam.sub.3CSK4 and R-MALP-2 lipopeptides
(EMC-Microcollections), or lipopolysaccharide (LPS, List
Biologicals) are dissolved in water to a concentration of 1 mg/mL,
sonicated 5 times for 30 seconds, and stored in aliquots at
-20.degree. C. Prior to addition to cells, an aliquot of the
dissolved ligand is sonicated for 1 minute and then is diluted in
medium to 2 ng/mL Pam3CSK4, 16 ng/mL R-MALP-2, or 100 ng/mL LPS.
The final concentration in the assay is 0.2 ng/mL Pam3CSK4, 1.6
ng/mL R-MALP-2, or 10 ng/mL LPS.
The test compounds are stored as 30 mM stocks in 4% DMSO. The final
concentrations of the test compounds in the assay are 0.1, 0.3, 1,
3, 10, and 30 .mu.M in 0.2% DMSO. 0.2% DMSO is used in the assay as
a control.
HEK293 cells stably carrying plasmids for TLR4, MD2, and the
NF-.kappa.B reporter gene ELAM-1-luciferase (HEK-TLR4-MD2-ELAM)
were generated as described by Yang et al., J. Biol. Chem.
275:20861-20866, 2000.
HEK-TLR2-ELAM cells were generated by a two-step method. In step 1,
Hek293 cells were transfected with pcDNA3.0 encoding human TLR2
followed by antibiotic selection with G418 in D-MEM supplemented
with 10% fetal bovine serum (Gibco BRL) to generate Hek-TLR2 cells.
In step 2, Hek-TLR2 cells were transfected with pELAM-luc/Zeo
followed by antibiotic selection with Zeocin in D-MEM supplemented
with 10% fetal bovine serum. Transfectants were screened for TLR2
responsiveness by measuring ligand-induced pELAM-luciferase
reporter activity, and analyzed for TLR2 mRNA expression by
RT-PCR.
TLR2 Antagonism Assay
Step 1. On day 1, HEK-TLR2-ELAM cells are plated at
2.times.10.sup.5 cells/mL, 80 .mu.L/well, in 96-well black plates.
The cells are incubated at 37.degree. C., 5% CO.sub.2, for 24
hours. The growth medium used is D-MEM, 10% fetal bovine serum, 2
mM L-glutamine, 10 .mu.g/mL Ciprofloxacin, 300 .mu.g/mL Geneticin
(G418), and 150 .mu.g/mL Zeocin.
Step 2. On day 2, 10 .mu.L of each test compound is added to the
wells, and 10 .mu.L lipopeptide is added to all of the wells. The
plates are then incubated at 37.degree. C., 5% CO.sub.2, for 18
hours.
Step 3. On day 3, 25 .mu.L of Steady-Glo reagent (Promega, Inc.) is
added to each well. The plates are then shaken for 5 minutes, and
the chemiluminescence of each well is read in a Wallac1450
MicroBetaTriLux counter. Dose-response curves were plotted in
KaleidaGraph, version 3.5 Synergy Software, and IC.sub.50 values
were calculated.
TLR4 Antagonism Assay
Step 1. On day 1, HEK-TLR4MD2-ELAM cells were plated at
4.times.10.sup.5 cells/mL, 80 .mu.L/well, in 96-well black plates.
The growth medium used is D-MEM, 10% fetal bovine serum, 2 mM
L-glutamine, 10 .mu.g/mL Ciprofloxacin, 300 .mu.g/mL Geneticin
(G418), 150 .mu.g/mL Zeocin, and 50 .mu.g/mL Hygromycin.
Step 2. On day 2, 10 .mu.L of each test compound is added to the
wells, and 10 .mu.L of LPS plus 10 nM soluble CD 14 (Biometec) is
added to all of the wells. The plates are then incubated at
37.degree. C., 5% CO.sub.2, for 18 hours.
Step 3. On day 3, 25 .mu.L of Steady-Glo reagent (Promega, Inc.) is
added to each well. The plates are then shaken for 5 minutes, and
the chemiluminescence of each well is read in a Wallac1450
MicroBetaTriLux counter. Dose-response curves were plotted in
KaleidaGraph, version 3.5 Synergy Software, and IC.sub.50 values
were calculated.
Antagonistic activities of selected compounds of the invention in
the TLR2 and TLR4 assays are presented in Table 6.
TABLE-US-00007 TABLE 6 TLR2 activity by Pam3CSK4 Compound
(IC.sub.50, .mu.M) TLR4 (IC.sub.50, .mu.M) ER811243 0.23 0.2
ER812011 0.24 6.6 ER811212 0.39 1.0 ER811245 0.43 3.4 ER811211 0.44
0.7 ER811393 0.46 12.3 ER811261 0.50 1.8 ER811395 0.65 1.8 ER811232
0.75 0.4 ER811254 0.77 3.6
TLR.sup.2 can cooperate with TLR.sup.6 and TLR.sup.1 to form
heterodimers and recognize different microbial ligands. For
example, TLR.sup.2 associates with TLR.sup.1 to recognize
triacylated lipopeptide (Pam3CSK4), but interacts with TLR.sup.6 to
recognize diacylated lipopeptide (R-MALP-2). Such heterodimer
selectivity for compounds of the invention is shown in Table 7 for
ER811245 and ER808977.
TABLE-US-00008 TABLE 7 TLR2 activity by TLR2 activity by Pam3CSK4
R-MALP-2 Compound (IC.sub.50, .mu.M) (IC.sub.50, .mu.M) ER-811245
0.44 30 ER-808977 0.24 20
From the foregoing description, it will be apparent that variations
and modifications may be made to the invention described herein to
adapt it to various usages and conditions. Such embodiments are
also within the scope of the following claims.
All publications mentioned in this specification are herein
incorporated by reference to the same extent as if each independent
publication or patent application was specifically and individually
indicated to be incorporated by reference.
Other embodiments are within the following claims.
* * * * *